S1 Supporting Information. Annotated vector sequences in GenBank format. Basic features, like polyadenylation signals, bacterial origin of replication, selection markers and CMV promoter were marked based on data found in Invitrogen pcDNA5/FRT/TO manual (Cat. no. V6520-20, Version G, 11 November 2010, 25-0368). >pKK-NoTag LOCUS Exported 5192 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-NoTag SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5192) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5192 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1150..1374 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1658..1705 /bound_moiety="FLP recombinase" /note="FRT" CDS 1713..2735 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2865..2995 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3378..4051) /direction=LEFT /note="pUC ori" CDS complement(4196..5056) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5057..5155) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatcc agcacagtgg cggccgctcg agtctagagg gcccgtttaa acccgctgat 1141 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt 1201 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat 1261 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg 1321 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg 1381 aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat 1441 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag 1501 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc 1561 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc 1621 ccaaaaaact tgattagggt gatggttcac gtacctagaa gttcctattc cgaagttcct 1681 attctctaga aagtatagga acttccttgg ccaaaaagcc tgaactcacc gcgacgtctg 1741 tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg 1801 gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa 1861 atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg 1921 cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca 1981 tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg 2041 ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga 2101 gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca 2161 tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca 2221 gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag 2281 tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca 2341 taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca 2401 acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc 2461 ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc cgcattggtc 2521 ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg 2581 gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc 2641 gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc 2701 gacgccccag cactcgtccg agggcaaagg aatagcacgt actacgagat ttcgattcca 2761 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga 2821 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag 2881 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt 2941 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac 3001 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt 3061 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg 3121 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt 3181 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt 3241 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc 3301 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg 3361 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg 3421 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac 3481 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg 3541 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct 3601 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg 3661 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct 3721 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac 3781 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt 3841 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc 3901 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca 3961 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat 4021 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac 4081 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt 4141 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc 4201 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg 4261 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg 4321 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc 4381 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta 4441 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg 4501 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct 4561 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta 4621 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg 4681 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga 4741 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt 4801 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca 4861 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt 4921 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt 4981 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga 5041 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt 5101 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc 5161 gcacatttcc ccgaaaagtg ccacctgacg tc // >pKK-FLAG-TEV LOCUS Exported 5213 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-FLAG-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5213) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5213 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1016 /codon_start=1 /product="FLAG" /note="FLAG" /translation="MDYKDDDDK" misc_feature 1023..1043 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1084..1104 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1171..1395 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1679..1726 /bound_moiety="FLP recombinase" /note="FRT" CDS 1734..2756 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2886..3016 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3399..4072) /direction=LEFT /note="pUC ori" CDS complement(4217..5077) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5078..5176) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tggattacaa ggatgacgac gataagggat 1021 ccgaaaacct gtacttccaa ggaaccggtg gtcacctgta caatcgatcc tgcaggagct 1081 agcgagaatt tgtattttca gggtgatatc cagcacagtg gcggccgctc gagtctagag 1141 ggcccgttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg 1201 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct 1261 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg 1321 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg 1381 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc 1441 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg 1501 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca 1561 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta 1621 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtacctaga 1681 agttcctatt ccgaagttcc tattctctag aaagtatagg aacttccttg gccaaaaagc 1741 ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac agcgtctccg 1801 acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc 1861 gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat cgttatgttt 1921 atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt ggggaattca 1981 gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg caagacctgc 2041 ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat gcgatcgctg 2101 cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga atcggtcaat 2161 acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa 2221 ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt 2281 gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg 2341 tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg atgttcgggg 2401 attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct tgtatggagc 2461 agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg cggctccggg 2521 cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac ggcaatttcg 2581 atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg 2641 tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag 2701 tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag gaatagcacg 2761 tactacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt 2821 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc 2881 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt 2941 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg 3001 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat 3061 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa 3121 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc 3181 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc 3241 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact 3301 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac 3361 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa 3421 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg 3481 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa 3541 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc 3601 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac 3661 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac 3721 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg 3781 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt 3841 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa 3901 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct 3961 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga 4021 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg 4081 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct 4141 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt 4201 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc 4261 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg 4321 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag 4381 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt 4441 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag 4501 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt 4561 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca 4621 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg 4681 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat 4741 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta 4801 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca 4861 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct 4921 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat 4981 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa 5041 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt 5101 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa 5161 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc // >pKK-HA-TEV LOCUS Exported 5216 bp ds-DNA circular SYN 13-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-HA-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5216) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5216 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1019 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /note="Met-HA" /translation="MYPYDVPDYA" misc_feature 1026..1046 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1087..1107 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1174..1398 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1682..1729 /bound_moiety="FLP recombinase" /note="FRT" CDS 1737..2759 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2889..3019 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3402..4075) /direction=LEFT /note="pUC ori" CDS complement(4220..5080) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5081..5179) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tgtaccccta cgacgtgccc gactacgctg 1021 gatccgaaaa cctgtacttc caaggaaccg gtggtcacct gtacaatcga tcctgcagga 1081 gctagcgaga atttgtattt tcagggtgat atccagcaca gtggcggccg ctcgagtcta 1141 gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg 1201 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt 1261 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg 1321 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg 1381 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc 1441 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 1501 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 1561 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 1621 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtacct 1681 agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc ttggccaaaa 1741 agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct 1801 ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag 1861 ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg 1921 tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat 1981 tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc 2041 tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg 2101 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc 2161 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc 2221 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc 2281 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca 2341 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg 2401 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg 2461 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc 2521 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt 2581 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga 2641 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag 2701 aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca aaggaatagc 2761 acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg 2821 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg 2881 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa 2941 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca 3001 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt 3061 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg 3121 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt 3181 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg 3241 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg 3301 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa 3361 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc 3421 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc 3481 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat 3541 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc 3601 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct 3661 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg 3721 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc 3781 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga 3841 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa 3901 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta 3961 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc 4021 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg 4081 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga 4141 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg 4201 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct 4261 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg 4321 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 4381 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa 4441 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 4501 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 4561 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 4621 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 4681 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 4741 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 4801 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 4861 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga 4921 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 4981 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 5041 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 5101 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga 5161 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc // >pKK-FLAG-BirA(R118G)-TEV LOCUS Exported 6182 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-FLAG-BirA(R118G)-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6182) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..6182 /organism="synthetic DNA construct" /mol_type="other DNA" source 1023..1985 /organism="Escherichia coli str. K-12 substr. MG1655" /strain="K-12" /sub_strain="MG1655" /mol_type="genomic DNA" /db_xref="taxon:511145" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1016 /codon_start=1 /product="FLAG" /note="FLAG" /translation="MDYKDDDDK" CDS 1023..1985 /codon_start=1 /transl_table=11 /gene="birA" /gene_synonym="bioR; dhbB; ECK3965; JW3941" /locus_tag="b3973" /product="bifunctional biotin-[acetylCoA carboxylase] holoenzyme synthetase/ DNA-binding transcriptional repressor, bio-5'-AMP-binding" /function="enzyme; Biosynthesis of cofactors, carriers: Biotin" /EC_number="6.3.4.15" /note="BirA(R118G)" /note="biotin-[acetylCoA carboxylase] holoenzyme synthetase and biotin operon repressor" /db_xref="GI:16131807" /db_xref="ASAP:ABE-0013001" /db_xref="UniProtKB/Swiss-Prot:P06709" /db_xref="EcoGene:EG10123" /db_xref="GeneID:948469" /protein_id="NP_418404.1" /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK" gene 1023..1985 /gene="birA" /gene_synonym="bioR; dhbB; ECK3965; JW3941" /locus_tag="b3973" /note="birA" /db_xref="EcoGene:EG10123" /db_xref="GeneID:948469" gene 1023..1026 /gene="murB" /gene_synonym="ECK3964; JW3940; yijB" /locus_tag="b3972" /note="murB" /db_xref="EcoGene:EG11205" /db_xref="GeneID:948470" misc_feature 1992..2012 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 2053..2073 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 2140..2364 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2648..2695 /bound_moiety="FLP recombinase" /note="FRT" CDS 2703..3725 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3855..3985 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4368..5041) /direction=LEFT /note="pUC ori" CDS complement(5186..6046) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(6047..6145) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggattacaa ggatgacgac gataagggat 1021 ccatgaagga taacaccgtg ccactgaaat tgattgccct gttagcgaac ggtgaatttc 1081 actctggcga gcagttgggt gaaacgctgg gaatgagccg ggcggctatt aataaacaca 1141 ttcagacact gcgtgactgg ggcgttgatg tctttaccgt tccgggtaaa ggatacagcc 1201 tgcctgagcc tatccagtta cttaatgcta aacagatatt gggtcagctg gatggcggta 1261 gtgtagccgt gctgccagtg attgactcca cgaatcagta ccttcttgat cgtatcggag 1321 agcttaaatc gggcgatgct tgcattgcag aataccagca ggctggccgt ggtcgccggg 1381 gtcggaaatg gttttcgcct tttggcgcaa acttatattt gtcgatgttc tggcgtctgg 1441 aacaaggccc ggcggcggcg attggtttaa gtctggttat cggtatcgtg atggcggaag 1501 tattacgcaa gctgggtgca gataaagttc gtgttaaatg gcctaatgac ctctatctgc 1561 aggatcgcaa gctggcaggc attctggtgg agctgactgg caaaactggc gatgcggcgc 1621 aaatagtcat tggagccggg atcaacatgg caatgcgccg tgttgaagag agtgtcgtta 1681 atcaggggtg gatcacgctg caggaagcgg ggatcaatct cgatcgtaat acgttggcgg 1741 ccatgctaat acgtgaatta cgtgctgcgt tggaactctt cgaacaagaa ggattggcac 1801 cttatctgtc gcgctgggaa aagctggata attttattaa tcgcccagtg aaacttatca 1861 ttggtgataa agaaatattt ggcatttcac gcggaataga caaacagggg gctttattac 1921 ttgagcagga tggaataata aaaccctgga tgggcggtga aatatccctg cgtagtgcag 1981 aaaaaggatc cgaaaacctg tacttccaag gaaccggtgg tcacctgtac aatcgatcct 2041 gcaggagcta gcgagaattt gtattttcag ggtgatatcc agcacagtgg cggccgctcg 2101 agtctagagg gcccgtttaa acccgctgat cagcctcgac tgtgccttct agttgccagc 2161 catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg 2221 tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc 2281 tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg 2341 ctggggatgc ggtgggctct atggcttctg aggcggaaag aaccagctgg ggctctaggg 2401 ggtatcccca cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca 2461 gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct 2521 ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt 2581 tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac 2641 gtacctagaa gttcctattc cgaagttcct attctctaga aagtatagga acttccttgg 2701 ccaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca 2761 gcgtctccga cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg 2821 taggagggcg tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc 2881 gttatgttta tcggcacttt gcatcggccg cgctcccgat tccggaagtg cttgacattg 2941 gggaattcag cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc 3001 aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg 3061 cgatcgctgc ggccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa 3121 tcggtcaata cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc 3181 actggcaaac tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc 3241 tgatgctttg ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct 3301 ccaacaatgt cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga 3361 tgttcgggga ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt 3421 gtatggagca gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc 3481 ggctccgggc gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg 3541 gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag 3601 ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc ggccgtctgg accgatggct 3661 gtgtagaagt actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg 3721 aatagcacgt actacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg 3781 gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt 3841 tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca 3901 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac 3961 tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat 4021 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac 4081 gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa 4141 ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat 4201 gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc 4261 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg 4321 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag 4381 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc 4441 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag 4501 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga 4561 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc 4621 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg 4681 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt 4741 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca 4801 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca 4861 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag 4921 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca 4981 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg 5041 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa 5101 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta 5161 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag 5221 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga 5281 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac 5341 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc 5401 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta 5461 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac 5521 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat 5581 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa 5641 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg 5701 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag 5761 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc 5821 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct 5881 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat 5941 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg 6001 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc 6061 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta 6121 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg 6181 tc // >pKK-MBP-TEV LOCUS Exported 6287 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-MBP-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6287) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..6287 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..2090 /codon_start=1 /gene="malE (mutated)" /product="maltose binding protein from E. coli" /note="MBP" /note="This version of the gene does not encode a signal sequence, so MBP will remain in the cytosol." /translation="MKTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF WYAVRTAVINAASGRQTVDEALKDAQT" misc_feature 2097..2117 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 2158..2178 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 2245..2469 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2753..2800 /bound_moiety="FLP recombinase" /note="FRT" CDS 2808..3830 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3960..4090 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4473..5146) /direction=LEFT /note="pUC ori" CDS complement(5291..6151) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(6152..6250) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tgaaaactga agaaggtaaa ctggtaatct 1021 ggattaacgg cgataaaggc tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag 1081 ataccggaat taaagtcacc gttgagcatc cggataaact ggaagagaaa ttcccacagg 1141 ttgcggcaac tggcgatggc cctgacatta tcttctgggc acacgaccgc tttggtggct 1201 acgctcaatc tggcctgttg gctgaaatca ccccggacaa agcgttccag gacaagctgt 1261 atccgtttac ctgggatgcc gtacgttaca acggcaagct gattgcttac ccgatcgctg 1321 ttgaagcgtt atcgctgatt tataacaaag atctgctgcc gaacccgcca aaaacctggg 1381 aagagatccc ggcgctggat aaagaactga aagcgaaagg taagagcgcg ctgatgttca 1441 acctgcaaga accgtacttc acctggccgc tgattgctgc tgacgggggt tatgcgttca 1501 agtatgaaaa cggcaagtac gacattaaag acgtgggcgt ggataacgct ggcgcgaaag 1561 cgggtctgac cttcctggtt gacctgatta aaaacaaaca catgaatgca gacaccgatt 1621 actccatcgc agaagctgcc tttaataaag gcgaaacagc gatgaccatc aacggcccgt 1681 gggcatggtc caacatcgac accagcaaag tgaattatgg tgtaacggta ctgccgacct 1741 tcaagggtca accatccaaa ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca 1801 gtccgaacaa agagctggcg aaagagttcc tcgaaaacta tctgctgact gatgaaggtc 1861 tggaagcggt taataaagac aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag 1921 agttggcgaa agatccacgt attgccgcca ccatggaaaa cgcccagaaa ggtgaaatca 1981 tgccgaacat cccgcagatg tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg 2041 ccgccagcgg tcgtcagact gtcgatgaag ccctgaaaga cgcgcagact ggatccgaaa 2101 acctgtactt ccaaggaacc ggtggtcacc tgtacaatcg atcctgcagg agctagcgag 2161 aatttgtatt ttcagggtga tatccagcac agtggcggcc gctcgagtct agagggcccg 2221 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc 2281 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa 2341 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg 2401 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg 2461 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc 2521 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac 2581 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg 2641 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt 2701 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtacc tagaagttcc 2761 tattccgaag ttcctattct ctagaaagta taggaacttc cttggccaaa aagcctgaac 2821 tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc tccgacctga 2881 tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga gggcgtggat 2941 atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa agatcgttat gtttatcggc 3001 actttgcatc ggccgcgctc ccgattccgg aagtgcttga cattggggaa ttcagcgaga 3061 gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa 3121 ccgaactgcc cgctgttctg cagccggtcg cggaggccat ggatgcgatc gctgcggccg 3181 atcttagcca gacgagcggg ttcggcccat tcggaccgca aggaatcggt caatacacta 3241 catggcgtga tttcatatgc gcgattgctg atccccatgt gtatcactgg caaactgtga 3301 tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg ctttgggccg 3361 aggactgccc cgaagtccgg cacctcgtgc acgcggattt cggctccaac aatgtcctga 3421 cggacaatgg ccgcataaca gcggtcattg actggagcga ggcgatgttc ggggattccc 3481 aatacgaggt cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga 3541 cgcgctactt cgagcggagg catccggagc ttgcaggatc gccgcggctc cgggcgtata 3601 tgctccgcat tggtcttgac caactctatc agagcttggt tgacggcaat ttcgatgatg 3661 cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc 3721 gtacacaaat cgcccgcaga agcgcggccg tctggaccga tggctgtgta gaagtactcg 3781 ccgatagtgg aaaccgacgc cccagcactc gtccgagggc aaaggaatag cacgtactac 3841 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg 3901 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca 3961 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa 4021 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt 4081 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt 4141 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa 4201 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac 4261 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg 4321 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc 4381 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat 4441 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca 4501 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc 4561 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc 4621 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg 4681 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta 4741 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg 4801 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac 4861 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag 4921 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat 4981 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat 5041 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc 5101 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt 5161 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct 5221 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt 5281 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc 5341 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac 5401 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat 5461 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg 5521 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata 5581 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta 5641 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt 5701 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag 5761 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa 5821 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc 5881 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt 5941 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc 6001 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta 6061 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa 6121 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca 6181 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac 6241 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc // >pKK-EGFP-TEV LOCUS Exported 5903 bp ds-DNA circular SYN 13-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-EGFP-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /product="enhanced GFP" /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 1081 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt 1201 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg 1441 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca 1501 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1681 ctctcggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc // >pKK-mCherry-TEV LOCUS Exported 5894 bp ds-DNA circular SYN 13-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mCherry-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5894) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5894 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1697 /codon_start=1 /note="mCherry" /note="mCherry" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" misc_feature 1704..1724 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1765..1785 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1852..2076 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2360..2407 /bound_moiety="FLP recombinase" /note="FRT" CDS 2415..3437 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3567..3697 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4080..4753) /direction=LEFT /note="pUC ori" CDS complement(4898..5758) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5759..5857) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag gataacatgg 1021 ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg 1081 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc 1141 tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca 1201 tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt 1261 ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga 1321 ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg 1381 gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct 1441 cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga 1501 agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc 1561 ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg 1621 aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca 1681 tggacgagct gtacaaggga tccgaaaacc tgtacttcca aggaaccggt ggtcacctgt 1741 acaatcgatc ctgcaggagc tagcgagaat ttgtattttc agggtgatat ccagcacagt 1801 ggcggccgct cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt 1861 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg 1921 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt 1981 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca 2041 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct 2101 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg 2161 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt 2221 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc 2281 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg 2341 gtgatggttc acgtacctag aagttcctat tccgaagttc ctattctcta gaaagtatag 2401 gaacttcctt ggccaaaaag cctgaactca ccgcgacgtc tgtcgagaag tttctgatcg 2461 aaaagttcga cagcgtctcc gacctgatgc agctctcgga gggcgaagaa tctcgtgctt 2521 tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt aaatagctgc gccgatggtt 2581 tctacaaaga tcgttatgtt tatcggcact ttgcatcggc cgcgctcccg attccggaag 2641 tgcttgacat tggggaattc agcgagagcc tgacctattg catctcccgc cgtgcacagg 2701 gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc tgttctgcag ccggtcgcgg 2761 aggccatgga tgcgatcgct gcggccgatc ttagccagac gagcgggttc ggcccattcg 2821 gaccgcaagg aatcggtcaa tacactacat ggcgtgattt catatgcgcg attgctgatc 2881 cccatgtgta tcactggcaa actgtgatgg acgacaccgt cagtgcgtcc gtcgcgcagg 2941 ctctcgatga gctgatgctt tgggccgagg actgccccga agtccggcac ctcgtgcacg 3001 cggatttcgg ctccaacaat gtcctgacgg acaatggccg cataacagcg gtcattgact 3061 ggagcgaggc gatgttcggg gattcccaat acgaggtcgc caacatcttc ttctggaggc 3121 cgtggttggc ttgtatggag cagcagacgc gctacttcga gcggaggcat ccggagcttg 3181 caggatcgcc gcggctccgg gcgtatatgc tccgcattgg tcttgaccaa ctctatcaga 3241 gcttggttga cggcaatttc gatgatgcag cttgggcgca gggtcgatgc gacgcaatcg 3301 tccgatccgg agccgggact gtcgggcgta cacaaatcgc ccgcagaagc gcggccgtct 3361 ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa ccgacgcccc agcactcgtc 3421 cgagggcaaa ggaatagcac gtactacgag atttcgattc caccgccgcc ttctatgaaa 3481 ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc 3541 tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat 3601 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg 3661 gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga 3721 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc 3781 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct 3841 aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc 3901 agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt 3961 ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag 4021 ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca 4081 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt 4141 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc 4201 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct 4261 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg 4321 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca 4381 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact 4441 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta 4501 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta 4561 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct 4621 tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt 4681 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga 4741 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca 4801 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat 4861 caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg 4921 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt 4981 agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag 5041 acccacgctc accggctcca gatttatcag caataaacca gccagccgga agggccgagc 5101 gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag 5161 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca 5221 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa 5281 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga 5341 tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata 5401 attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca 5461 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg 5521 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg 5581 ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg 5641 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag 5701 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac 5761 tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca 5821 tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag 5881 tgccacctga cgtc // >pKK-mCerulean-TEV LOCUS Exported 5903 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mCerulean-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /note="mCerulean" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 1081 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctgg ggcgtgcagt 1201 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caacgccatc agcgacaacg 1441 tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca 1501 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1681 ctctcggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc // >pKK-mAmber-TEV LOCUS Exported 5903 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mAmber-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /product="mVenus YFP with monomerizing A206K mutation (Nagai et al., 2002)" /note="mAmber" /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGCGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 1081 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagctc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctcggctgc ggcctgcagt 1201 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg 1441 tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca 1501 acatcgagga cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccaag ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1681 ctctcggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc // >pKK-CyOFP1-TEV LOCUS Exported 5888 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-CyOFP1-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5888) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5888 /organism="synthetic DNA construct" /mol_type="other DNA" source 990..1691 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1691 /codon_start=1 /note="CyOFP1" /translation="MVSKGEELIKENMRSKLYLEGSVNGHQFKCTHEGEGKPYEGKQTN RIKVVEGGPLPFAFDILATHFMYGSKVFIKYPADLPDYFKQSFPEGFTWERVMVFEDGG VLTATQDTSLQDGELIYNVKVRGVNFPANGPVMQKKTLGWEPSTETMYPADGGLEGRCD KALKLVGGGHLHVNFKTTYKSKKPVKMPGVHYVDRRLERIKEADNETYVEQYEHAVARY SNLGGGMDELYK" misc_feature 1698..1718 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1759..1779 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1846..2070 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2354..2401 /bound_moiety="FLP recombinase" /note="FRT" CDS 2409..3431 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3561..3691 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4074..4747) /direction=LEFT /note="pUC ori" CDS complement(4892..5752) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5753..5851) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgatcaagg 1021 agaacatgag aagcaagctg tacctggaag gcagcgtgaa cggccaccag ttcaagtgca 1081 cccacgaagg ggagggcaag ccctacgagg gcaagcagac caacaggatc aaggtggtgg 1141 agggaggccc cctgccgttc gcattcgaca tcctggccac ccactttatg tacgggagca 1201 aggtgttcat caagtacccc gccgacctcc ccgattattt taagcagtcc ttccctgagg 1261 gcttcacatg ggagagagtc atggtgttcg aagacggggg cgtgctgacc gccacccagg 1321 acaccagcct ccaggacggc gagctcatct acaacgtcaa ggtcagaggg gtgaacttcc 1381 cagccaacgg ccccgtgatg cagaagaaaa cactgggctg ggagcccagc accgagacca 1441 tgtaccccgc tgacggcggc ctggaaggca gatgcgacaa ggccctgaag ctcgtgggcg 1501 ggggccacct gcacgtcaac ttcaagacca catacaagtc caagaaaccc gtgaagatgc 1561 ccggcgtcca ctacgtggac cgcagactgg aaagaatcaa ggaggccgac aacgagacct 1621 acgtcgagca gtacgagcac gctgtggcca gatactccaa cctgggcgga ggcatggacg 1681 agctgtacaa gggatccgaa aacctgtact tccaaggaac cggtggtcac ctgtacaatc 1741 gatcctgcag gagctagcga gaatttgtat tttcagggtg atatccagca cagtggcggc 1801 cgctcgagtc tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt 1861 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc 1921 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt 1981 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca 2041 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct 2101 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta 2161 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc 2221 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt 2281 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg 2341 gttcacgtac ctagaagttc ctattccgaa gttcctattc tctagaaagt ataggaactt 2401 ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt 2461 tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct 2521 tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca 2581 aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg 2641 acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca 2701 cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca 2761 tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc 2821 aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg 2881 tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg 2941 atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt 3001 tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg 3061 aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt 3121 tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat 3181 cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg 3241 ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat 3301 ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg 3361 atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg 3421 caaaggaata gcacgtacta cgagatttcg attccaccgc cgccttctat gaaaggttgg 3481 gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc 3541 tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca 3601 atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt 3661 ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg 3721 cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca 3781 acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca 3841 cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc 3901 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt 3961 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact 4021 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag 4081 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata 4141 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc 4201 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg 4261 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc 4321 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg 4381 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc 4441 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga 4501 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg 4561 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa 4621 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg 4681 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt 4741 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat 4801 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct 4861 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta 4921 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa 4981 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac 5041 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa 5101 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag 5161 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg 5221 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag 5281 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg 5341 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc 5401 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat 5461 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata 5521 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa 5581 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca 5641 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc 5701 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc 5761 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg 5821 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac 5881 ctgacgtc // >pKK-Dendra2N-TEV LOCUS Exported 5876 bp ds-DNA circular SYN 27-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-Dendra2N-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5876) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5876 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1679 /codon_start=1 /product="monomeric photoswitchable green-to-red fluorescent protein from Dendronephthya" /note="Dendra2N" /note="mammalian codon-optimized" /translation="MNTPGINLIKEDMRVKVHMEGNVNGHAFVIEGEGKGKPYEGTQTA NLTVKEGAPLPFSYDILTTAVHYGNRVFTKYPEDIPDYFKQSFPEGYSWERTMTFEDKG ICTIRSDISLEGDCFFQNVRFKGTNFPPNGPVMQKKTLKWEPSTEKLHVRDGLLVGNIN MALLLEGGGHYLCDFKTTYKAKKVVQLPDAHFVDHRIEILGNDSDYNKVKLYEHAVARY SPLPSQVW" misc_feature 1686..1706 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1747..1767 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1834..2058 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2342..2389 /bound_moiety="FLP recombinase" /note="FRT" CDS 2397..3419 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3549..3679 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4062..4735) /direction=LEFT /note="pUC ori" CDS complement(4880..5740) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5741..5839) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tgaacacccc gggaattaac ctgatcaagg 1021 aggacatgcg cgtgaaggtg cacatggagg gcaacgtgaa cggccacgcc ttcgtgatcg 1081 agggcgaggg caagggcaag ccctacgagg gcacccagac cgccaacctg accgtgaagg 1141 agggcgcccc cctgcccttc agctacgaca tcctgaccac cgccgtgcac tacggcaacc 1201 gggtgttcac caagtacccc gaggacatcc ccgactactt caagcagagc ttccccgagg 1261 gctacagctg ggagcgcacc atgaccttcg aggacaaggg catctgcacc atccgcagcg 1321 acatcagcct ggagggcgac tgcttcttcc agaacgtgcg cttcaagggc accaacttcc 1381 cccccaacgg ccccgtgatg cagaagaaga ccctgaagtg ggagcccagc accgagaagc 1441 tgcacgtgcg cgacggcctg ctggtgggca acatcaacat ggccctgctg ctggagggcg 1501 gcggccacta cctgtgcgac ttcaagacca cctacaaggc caagaaggtg gtgcagctgc 1561 ccgacgccca cttcgtggac caccgcatcg agatcctggg caacgacagc gactacaaca 1621 aggtgaagct gtacgagcac gccgtggccc gctacagccc cctgcccagc caggtgtggg 1681 gatccgaaaa cctgtacttc caaggaaccg gtggtcacct gtacaatcga tcctgcagga 1741 gctagcgaga atttgtattt tcagggtgat atccagcaca gtggcggccg ctcgagtcta 1801 gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg 1861 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt 1921 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg 1981 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg 2041 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc 2101 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 2161 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 2221 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 2281 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtacct 2341 agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc ttggccaaaa 2401 agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct 2461 ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag 2521 ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg 2581 tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat 2641 tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc 2701 tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg 2761 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc 2821 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc 2881 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc 2941 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca 3001 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg 3061 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg 3121 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc 3181 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt 3241 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga 3301 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag 3361 aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca aaggaatagc 3421 acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg 3481 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg 3541 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa 3601 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca 3661 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt 3721 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg 3781 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt 3841 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg 3901 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg 3961 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa 4021 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc 4081 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc 4141 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat 4201 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc 4261 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct 4321 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg 4381 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc 4441 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga 4501 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa 4561 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta 4621 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc 4681 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg 4741 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga 4801 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg 4861 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct 4921 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg 4981 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 5041 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa 5101 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 5161 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 5221 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 5281 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 5341 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 5401 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 5461 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 5521 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga 5581 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 5641 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 5701 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 5761 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga 5821 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc // >pKK-mCardinal-TEV LOCUS Exported 5918 bp ds-DNA circular SYN 23-NOV-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mCardinal-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5918) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Monday, May 22, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5918 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1721 /codon_start=1 /product="far-red fluorescent protein derived from mNeptune, with further red-shifted spectra" /note="mCardinal" /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTTEGEGKPYEGTQTQ RIKVVEGGPLPFAFDILATCFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG VLTVTQDTSLQDGCLIYNVKLRGVNFPSNGPVMQKKTLGWEATTETLYPADGGLEGRCD MALKLVGGGHLHCNLKTTYRSKKPAKNLKMPGVYFVDRRLERIKEADNETYVEQHEVAV ARYCDLPSKLGHKLNGMDELYK" misc_feature 1728..1748 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1789..1809 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1876..2100 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2384..2431 /bound_moiety="FLP recombinase" /note="FRT" CDS 2439..3461 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3591..3721 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4104..4777) /direction=LEFT /note="pUC ori" CDS complement(4922..5782) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5783..5881) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgatcaagg 1021 agaacatgca catgaagctg tacatggaag gcaccgtgaa caaccaccac ttcaagtgca 1081 ccaccgaagg ggagggcaag ccctacgagg gcacccagac ccagaggatt aaggtggtgg 1141 agggaggccc cctgccgttc gcattcgaca tcctggccac ctgctttatg tacgggagca 1201 agaccttcat caaccacacc cagggcatcc ccgatttctt taagcagtcc ttccctgagg 1261 gcttcacatg ggagagagtc accacatacg aagacggggg cgtgcttacc gttacccagg 1321 acaccagcct ccaggacggc tgcttgatct acaacgtcaa gctcagaggg gtgaacttcc 1381 catccaacgg ccctgtgatg cagaagaaaa cactcggctg ggaggccacc accgagaccc 1441 tgtaccccgc tgacggcggc ctggaaggca gatgcgacat ggccctgaag ctcgtgggcg 1501 ggggccacct gcactgcaac ctgaagacca catacagatc caagaaaccc gctaagaacc 1561 tcaagatgcc cggcgtctac tttgtggacc gcagactgga aagaatcaag gaggccgaca 1621 atgagaccta cgtcgagcag cacgaggtgg ctgtggccag atactgcgac ctccctagca 1681 aactggggca caaacttaat ggcatggacg agctgtacaa gggatccgaa aacctgtact 1741 tccaaggaac cggtggtcac ctgtacaatc gatcctgcag gagctagcga gaatttgtat 1801 tttcagggtg atatccagca cagtggcggc cgctcgagtc tagagggccc gtttaaaccc 1861 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg 1921 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa 1981 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca 2041 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg 2101 cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg 2161 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg 2221 ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc 2281 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc 2341 tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc ctattccgaa 2401 gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa ctcaccgcga 2461 cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg atgcagctct 2521 cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga tatgtcctgc 2581 gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg cactttgcat 2641 cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag agcctgacct 2701 attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa accgaactgc 2761 ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc gatcttagcc 2821 agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact acatggcgtg 2881 atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg atggacgaca 2941 ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc gaggactgcc 3001 ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg acggacaatg 3061 gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc caatacgagg 3121 tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag acgcgctact 3181 tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat atgctccgca 3241 ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat gcagcttggg 3301 cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg cgtacacaaa 3361 tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc gccgatagtg 3421 gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtacta cgagatttcg 3481 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct 3541 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta 3601 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat 3661 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct 3721 gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt 3781 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag 3841 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt 3901 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag 3961 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg 4021 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat 4081 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta 4141 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa 4201 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc 4261 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt 4321 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca 4381 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg 4441 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat 4501 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta 4561 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct 4621 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac 4681 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa 4741 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa 4801 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt 4861 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca 4921 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca 4981 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc 5041 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa 5101 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc 5161 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca 5221 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat 5281 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag 5341 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac 5401 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt 5461 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt 5521 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc 5581 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat 5641 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca 5701 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga 5761 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg 5821 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg 5881 ttccgcgcac atttccccga aaagtgccac ctgacgtc // >pKK-mClover3-TEV LOCUS Exported 5903 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mClover3-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" source 990..1706 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /note="mClover3" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtcc 1081 gcggcgaggg cgagggcgat gccaccaacg gcaagctgac cctgaagttc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cttcggctac ggcgtggcct 1201 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatctctt tcaaggacga cggtacctac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caacttcaac agccactacg 1441 tctatatcac ggccgacaag cagaagaact gcatcaaggc taacttcaag atccgccaca 1501 acgttgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcca tcagtccaag ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatta 1681 cacatggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc // >pKK-mRuby3-TEV LOCUS Exported 5897 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mRuby3-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5897) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5897 /organism="synthetic DNA construct" /mol_type="other DNA" source 990..1700 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1700 /codon_start=1 /note="mRuby3" /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV AKYSNLGGGMDELYK" misc_feature 1707..1727 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1768..1788 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1855..2079 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2363..2410 /bound_moiety="FLP recombinase" /note="FRT" CDS 2418..3440 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3570..3700 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4083..4756) /direction=LEFT /note="pUC ori" CDS complement(4901..5761) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5762..5860) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgtctaa gggcgaagag ctgatcaagg 1021 aaaatatgcg tatgaaggtg gtcatggaag gttcggtcaa cggccaccaa ttcaaatgca 1081 caggtgaagg agaaggcaga ccgtacgagg gagtgcaaac catgaggatc aaagtcatcg 1141 agggaggacc cctgccattt gcctttgaca ttcttgccac gtcgttcatg tatggcagcc 1201 gtacctttat caagtacccg gccgacatcc ctgatttctt taaacagtcc tttcctgagg 1261 gttttacttg ggaaagagtt acgagatacg aagatggtgg agtcgtcacc gtcacgcagg 1321 acaccagcct tgaggatggc gagctcgtct acaacgtcaa ggtcagaggg gtaaactttc 1381 cctccaatgg tcccgtgatg cagaagaaga ccaagggttg ggagcctaat acagagatga 1441 tgtatccagc agatggtggt ctgagaggat acactgacat cgcactgaaa gttgatggtg 1501 gtggccatct gcactgcaac ttcgtgacaa cttacaggtc aaaaaagacc gtcgggaaca 1561 tcaagatgcc cggtgtccat gccgttgatc accgcctgga aaggatcgag gagagtgaca 1621 atgaaaccta cgtagtgcaa agagaagtgg cagttgccaa atacagcaac cttggtggtg 1681 gcatggacga gctgtacaag ggatccgaaa acctgtactt ccaaggaacc ggtggtcacc 1741 tgtacaatcg atcctgcagg agctagcgag aatttgtatt ttcagggtga tatccagcac 1801 agtggcggcc gctcgagtct agagggcccg tttaaacccg ctgatcagcc tcgactgtgc 1861 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag 1921 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta 1981 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag 2041 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca 2101 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg 2161 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg 2221 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg 2281 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt 2341 agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct ctagaaagta 2401 taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag aagtttctga 2461 tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa gaatctcgtg 2521 ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg 2581 gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc ccgattccgg 2641 aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac 2701 agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg 2761 cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg ttcggcccat 2821 tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc gcgattgctg 2881 atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc 2941 aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg cacctcgtgc 3001 acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca gcggtcattg 3061 actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc ttcttctgga 3121 ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg catccggagc 3181 ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc 3241 agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa 3301 tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg 3361 tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc 3421 gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg 3481 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg 3541 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca 3601 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt 3661 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct 3721 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa 3781 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga 3841 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt 3901 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct 3961 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat 4021 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga 4081 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt 4141 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt 4201 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc 4261 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa 4321 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct 4381 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta 4441 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg 4501 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc 4561 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta 4621 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg 4681 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt 4741 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg 4801 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta 4861 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg 4921 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg 4981 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc 5041 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg 5101 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg 5161 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag 5221 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat 5281 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc 5341 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc 5401 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa 5461 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac 5521 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt 5581 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc 5641 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa 5701 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca 5761 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat 5821 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa 5881 aagtgccacc tgacgtc // >pKK-mTagBFP-TEV LOCUS Exported 5885 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mTagBFP-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5885) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5885 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1688 /codon_start=1 /product="monomeric blue fluorescent protein" /note="mTagBFP" /note="mammalian codon-optimized" /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC DLPSKLGHKLN" misc_feature 1695..1715 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1756..1776 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1843..2067 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2351..2398 /bound_moiety="FLP recombinase" /note="FRT" CDS 2406..3428 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3558..3688 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4071..4744) /direction=LEFT /note="pUC ori" CDS complement(4889..5749) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5750..5848) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtacca tgagcgagct gattaaggag aacatgcaca 1021 tgaagctgta catggagggc accgtggaca accatcactt caagtgcaca tccgagggcg 1081 aaggcaagcc ctacgagggc acccagacca tgagaatcaa ggtggtcgag ggcggccctc 1141 tccccttcgc cttcgacatc ctggctacta gcttcctcta cggcagcaag accttcatca 1201 accacaccca gggcatcccc gacttcttca agcagtcctt ccctgagggc ttcacatggg 1261 agagagtcac cacatacgaa gacgggggcg tgctgaccgc tacccaggac accagcctcc 1321 aggacggctg cctcatctac aacgtcaaga tcagaggggt gaacttcaca tccaacggcc 1381 ctgtgatgca gaagaaaaca ctcggctggg aggccttcac cgagacgctg taccccgctg 1441 acggcggcct ggaaggcaga aacgacatgg ccctgaagct cgtgggcggg agccatctga 1501 tcgcaaacat caagaccaca tatagatcca agaaacccgc taagaacctc aagatgcctg 1561 gcgtctacta tgtggactac agactggaaa gaatcaagga ggccaacaac gagacctacg 1621 tcgagcagca cgaggtggca gtggccagat actgcgacct ccctagcaaa ctggggcaca 1681 agcttaatgg atccgaaaac ctgtacttcc aaggaaccgg tggtcacctg tacaatcgat 1741 cctgcaggag ctagcgagaa tttgtatttt cagggtgata tccagcacag tggcggccgc 1801 tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc 1861 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca 1921 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta 1981 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc 2041 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta 2101 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc 2161 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt 2221 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag 2281 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt 2341 cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata ggaacttcct 2401 tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg 2461 acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg 2521 atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag 2581 atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca 2641 ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt 2701 tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggccatgg 2761 atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag 2821 gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt 2881 atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg 2941 agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg 3001 gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg 3061 cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg 3121 cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc 3181 cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg 3241 acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg 3301 gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg 3361 gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt ccgagggcaa 3421 aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct 3481 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg 3541 agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata 3601 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca 3661 aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt 3721 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca 3781 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat 3841 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt 3901 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct 3961 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa 4021 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa 4081 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc 4141 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga 4201 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc 4261 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt 4321 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct 4381 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg 4441 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta 4501 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct 4561 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa 4621 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt 4681 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta 4741 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat 4801 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa 4861 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct 4921 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta 4981 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct 5041 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg 5101 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa 5161 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt 5221 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta 5281 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca 5341 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta 5401 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct 5461 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg 5521 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac 5581 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact 5641 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa 5701 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt 5761 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat 5821 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg 5881 acgtc // >pKK-mVenus-TEV LOCUS Exported 5903 bp ds-DNA circular SYN 26-OCT-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mVenus-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /product="Venus YFP with monomerizing A206K mutation (Nagai et al., 2002)" /note="mVenus" /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 1081 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagctc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctcggctac ggcctgcagt 1201 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg 1441 tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag atccgccaca 1501 acatcgagga cggcggcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccaag ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1681 ctctcggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc // >pKK-mEGFP-TEV LOCUS Exported 5903 bp ds-DNA circular SYN 09-MAR-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-mEGFP-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5903) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Monday, May 22, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5903 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1706 /codon_start=1 /product="enhanced GFP" /note="mEGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 1713..1733 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1774..1794 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1861..2085 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2369..2416 /bound_moiety="FLP recombinase" /note="FRT" CDS 2424..3446 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3576..3706 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4089..4762) /direction=LEFT /note="pUC ori" CDS complement(4907..5767) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5768..5866) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggtgagcaa gggcgaggag ctgttcaccg 1021 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 1081 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 1141 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt 1201 gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 1261 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 1321 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 1381 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg 1441 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca 1501 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 1561 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag ctgagcaaag 1621 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 1681 ctctcggcat ggacgagctg tacaagggat ccgaaaacct gtacttccaa ggaaccggtg 1741 gtcacctgta caatcgatcc tgcaggagct agcgagaatt tgtattttca gggtgatatc 1801 cagcacagtg gcggccgctc gagtctagag ggcccgttta aacccgctga tcagcctcga 1861 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc 1921 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc 1981 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt 2041 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa 2101 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg 2161 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc 2221 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa 2281 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac 2341 ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc tattctctag 2401 aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt 2461 ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag ggcgaagaat 2521 ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta aatagctgcg 2581 ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc gcgctcccga 2641 ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc atctcccgcc 2701 gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct gttctgcagc 2761 cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg agcgggttcg 2821 gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc atatgcgcga 2881 ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg 2941 tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa gtccggcacc 3001 tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc ataacagcgg 3061 tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc aacatcttct 3121 tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag cggaggcatc 3181 cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt cttgaccaac 3241 tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg 3301 acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg 3361 cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac cgacgcccca 3421 gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc accgccgcct 3481 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc 3541 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg 3601 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt 3661 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct 3721 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc 3781 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat 3841 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc 3901 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg 3961 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag 4021 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag 4081 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 4141 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 4201 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 4261 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 4321 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 4381 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 4441 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 4501 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 4561 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 4621 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 4681 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 4741 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 4801 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 4861 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 4921 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc 4981 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga 5041 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa 5101 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt 5161 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg 5221 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc 5281 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg 5341 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag 5401 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt 5461 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt 5521 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac 5581 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac 5641 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag 5701 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa 5761 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga 5821 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc 5881 cccgaaaagt gccacctgac gtc // >pKK-TEV-FLAG LOCUS Exported 5202 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-FLAG SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5202) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5202 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..1116 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1116 /codon_start=1 /note="FLAG" /translation="DYKDDDDK" polyA_signal 1160..1384 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1668..1715 /bound_moiety="FLP recombinase" /note="FRT" CDS 1723..2745 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2875..3005 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3388..4061) /direction=LEFT /note="pUC ori" CDS complement(4206..5066) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5067..5165) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatcg attacaagga tgacgacgat aagtagctcg agtctagagg gcccgtttaa 1141 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc 1201 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag 1261 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag 1321 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct 1381 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt 1441 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc 1501 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc 1561 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg 1621 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtacctagaa gttcctattc 1681 cgaagttcct attctctaga aagtatagga acttccttgg ccaaaaagcc tgaactcacc 1741 gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag 1801 ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc 1861 ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt 1921 gcatcggccg cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg 1981 acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa 2041 ctgcccgctg ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt 2101 agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg 2161 cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac 2221 gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac 2281 tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac 2341 aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac 2401 gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc 2461 tacttcgagc ggaggcatcc ggagcttgca ggatcgccgc ggctccgggc gtatatgctc 2521 cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga tgatgcagct 2581 tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca 2641 caaatcgccc gcagaagcgc ggccgtctgg accgatggct gtgtagaagt actcgccgat 2701 agtggaaacc gacgccccag cactcgtccg agggcaaagg aatagcacgt actacgagat 2761 ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc 2821 ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg 2881 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa 2941 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat 3001 gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct 3061 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt 3121 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc 3181 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg 3241 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg 3301 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca 3361 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac 3421 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac 3481 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg 3541 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac 3601 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat 3661 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag 3721 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac 3781 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt 3841 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt 3901 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc 3961 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga 4021 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac 4081 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc 4141 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct 4201 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca 4261 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct 4321 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca 4381 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc 4441 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg 4501 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct 4561 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa 4621 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta 4681 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc 4741 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg 4801 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa 4861 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg 4921 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc 4981 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg 5041 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat 5101 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata 5161 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc // >pKK-TEV-3XFLAG LOCUS Exported 5250 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-3XFLAG SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5250) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5250 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1108..1173 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /note="3xFLAG" /translation="DYKDHDGDYKDHDIDYKDDDDK" polyA_signal 1208..1432 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1716..1763 /bound_moiety="FLP recombinase" /note="FRT" CDS 1771..2793 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2923..3053 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3436..4109) /direction=LEFT /note="pUC ori" CDS complement(4254..5114) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5115..5213) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatcc agcacagtgg cggccgcgac tacaaagacc atgacggtga ttataaagat 1141 catgacatcg attacaagga tgacgatgac aagtaagggc ccgtttaaac ccgctgatca 1201 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc 1261 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg 1321 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg 1381 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag 1441 gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta 1501 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg 1561 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa 1621 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc 1681 aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat 1741 tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc 1801 gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc 1861 gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat 1921 agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg 1981 ctcccgattc cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc 2041 tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt 2101 ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc 2161 gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata 2221 tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt 2281 gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc 2341 cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata 2401 acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac 2461 atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg 2521 aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt 2581 gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt 2641 cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc 2701 agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga 2761 cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc 2821 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc 2881 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct 2941 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca 3001 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg 3061 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt 3121 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt 3181 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg 3241 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg 3301 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 3361 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 3421 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 3481 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 3541 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga 3601 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt 3661 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg 3721 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc 3781 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg 3841 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc 3901 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg 3961 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc 4021 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct 4081 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt 4141 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa 4201 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa 4261 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc 4321 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct 4381 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca 4441 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt 4501 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt 4561 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc 4621 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc 4681 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt 4741 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact 4801 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc 4861 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt 4921 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg 4981 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct 5041 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa 5101 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt 5161 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc 5221 acatttcccc gaaaagtgcc acctgacgtc // >pKK-TEV-HA LOCUS Exported 5205 bp ds-DNA circular SYN 13-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-HA SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5205) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5205 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1116 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /note="HA" /translation="YPYDVPDYA" polyA_signal 1163..1387 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1671..1718 /bound_moiety="FLP recombinase" /note="FRT" CDS 1726..2748 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2878..3008 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3391..4064) /direction=LEFT /note="pUC ori" CDS complement(4209..5069) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5070..5168) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatct acccctacga cgtgcccgac tacgcttgac tcgagtctag agggcccgtt 1141 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc 1201 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat 1261 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg 1321 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc 1381 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc 1441 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt 1501 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc 1561 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta 1621 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtaccta gaagttccta 1681 ttccgaagtt cctattctct agaaagtata ggaacttcct tggccaaaaa gcctgaactc 1741 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg 1801 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat 1861 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac 1921 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc 1981 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc 2041 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat 2101 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca 2161 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg 2221 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag 2281 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg 2341 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa 2401 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg 2461 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg 2521 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca 2581 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt 2641 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc 2701 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtactacga 2761 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac 2821 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac 2881 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat 2941 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat 3001 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt 3061 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag 3121 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg 3181 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg 3241 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc 3301 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc 3361 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg 3421 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat 3481 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag 3541 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga 3601 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg 3661 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt 3721 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac 3781 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc 3841 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt 3901 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc 3961 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc 4021 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg 4081 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag 4141 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg 4201 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt 4261 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca 4321 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca 4381 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc 4441 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt 4501 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg 4561 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc 4621 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg 4681 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga 4741 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga 4801 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta 4861 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg 4921 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact 4981 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata 5041 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt 5101 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa 5161 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtc // >pKK-TEV-MBP LOCUS Exported 6279 bp ds-DNA circular SYN 13-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-MBP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6279) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..6279 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..2190 /codon_start=1 /gene="malE (mutated)" /product="maltose binding protein from E. coli" /note="MBP" /note="This version of the gene does not encode a signal sequence, so MBP will remain in the cytosol." /translation="MKTEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF WYAVRTAVINAASGRQTVDEALKDAQT" polyA_signal 2237..2461 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2745..2792 /bound_moiety="FLP recombinase" /note="FRT" CDS 2800..3822 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3952..4082 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4465..5138) /direction=LEFT /note="pUC ori" CDS complement(5283..6143) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(6144..6242) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tgaaaactga agaaggtaaa ctggtaatct ggattaacgg cgataaaggc 1141 tataacggtc tcgctgaagt cggtaagaaa ttcgagaaag ataccggaat taaagtcacc 1201 gttgagcatc cggataaact ggaagagaaa ttcccacagg ttgcggcaac tggcgatggc 1261 cctgacatta tcttctgggc acacgaccgc tttggtggct acgctcaatc tggcctgttg 1321 gctgaaatca ccccggacaa agcgttccag gacaagctgt atccgtttac ctgggatgcc 1381 gtacgttaca acggcaagct gattgcttac ccgatcgctg ttgaagcgtt atcgctgatt 1441 tataacaaag atctgctgcc gaacccgcca aaaacctggg aagagatccc ggcgctggat 1501 aaagaactga aagcgaaagg taagagcgcg ctgatgttca acctgcaaga accgtacttc 1561 acctggccgc tgattgctgc tgacgggggt tatgcgttca agtatgaaaa cggcaagtac 1621 gacattaaag acgtgggcgt ggataacgct ggcgcgaaag cgggtctgac cttcctggtt 1681 gacctgatta aaaacaaaca catgaatgca gacaccgatt actccatcgc agaagctgcc 1741 tttaataaag gcgaaacagc gatgaccatc aacggcccgt gggcatggtc caacatcgac 1801 accagcaaag tgaattatgg tgtaacggta ctgccgacct tcaagggtca accatccaaa 1861 ccgttcgttg gcgtgctgag cgcaggtatt aacgccgcca gtccgaacaa agagctggcg 1921 aaagagttcc tcgaaaacta tctgctgact gatgaaggtc tggaagcggt taataaagac 1981 aaaccgctgg gtgccgtagc gctgaagtct tacgaggaag agttggcgaa agatccacgt 2041 attgccgcca ccatggaaaa cgcccagaaa ggtgaaatca tgccgaacat cccgcagatg 2101 tccgctttct ggtatgccgt gcgtactgcg gtgatcaacg ccgccagcgg tcgtcagact 2161 gtcgatgaag ccctgaaaga cgcgcagact taactcgagt ctagagggcc cgtttaaacc 2221 cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc 2281 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa 2341 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac 2401 agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg 2461 gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc 2521 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc 2581 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt 2641 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac 2701 ctcgacccca aaaaacttga ttagggtgat ggttcacgta cctagaagtt cctattccga 2761 agttcctatt ctctagaaag tataggaact tccttggcca aaaagcctga actcaccgcg 2821 acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg tctccgacct gatgcagctc 2881 tcggagggcg aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg atatgtcctg 2941 cgggtaaata gctgcgccga tggtttctac aaagatcgtt atgtttatcg gcactttgca 3001 tcggccgcgc tcccgattcc ggaagtgctt gacattgggg aattcagcga gagcctgacc 3061 tattgcatct cccgccgtgc acagggtgtc acgttgcaag acctgcctga aaccgaactg 3121 cccgctgttc tgcagccggt cgcggaggcc atggatgcga tcgctgcggc cgatcttagc 3181 cagacgagcg ggttcggccc attcggaccg caaggaatcg gtcaatacac tacatggcgt 3241 gatttcatat gcgcgattgc tgatccccat gtgtatcact ggcaaactgt gatggacgac 3301 accgtcagtg cgtccgtcgc gcaggctctc gatgagctga tgctttgggc cgaggactgc 3361 cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca acaatgtcct gacggacaat 3421 ggccgcataa cagcggtcat tgactggagc gaggcgatgt tcggggattc ccaatacgag 3481 gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta tggagcagca gacgcgctac 3541 ttcgagcgga ggcatccgga gcttgcagga tcgccgcggc tccgggcgta tatgctccgc 3601 attggtcttg accaactcta tcagagcttg gttgacggca atttcgatga tgcagcttgg 3661 gcgcagggtc gatgcgacgc aatcgtccga tccggagccg ggactgtcgg gcgtacacaa 3721 atcgcccgca gaagcgcggc cgtctggacc gatggctgtg tagaagtact cgccgatagt 3781 ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat agcacgtact acgagatttc 3841 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc 3901 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt 3961 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca 4021 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc 4081 tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg 4141 tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa 4201 gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct 4261 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga 4321 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc 4381 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa 4441 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt 4501 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa 4561 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt 4621 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg 4681 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc 4741 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc 4801 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta 4861 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct 4921 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc 4981 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa 5041 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa 5101 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa 5161 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt 5221 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac 5281 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc 5341 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc 5401 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata 5461 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc 5521 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc 5581 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca 5641 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa 5701 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca 5761 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt 5821 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt 5881 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg 5941 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga 6001 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc 6061 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg 6121 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag 6181 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg 6241 gttccgcgca catttccccg aaaagtgcca cctgacgtc // >pKK-TEV-ProteinA LOCUS Exported 5589 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-ProteinA SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5589) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5589 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1503 /codon_start=1 /note="protein A" /translation="KTAALAQHDEAVDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSL KDDPSQSANLLAEAKKLNDAQAPKVDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLK DDPSQSANLLAEAKKLNGAQAPKVDANSAGKST" polyA_signal 1547..1771 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2055..2102 /bound_moiety="FLP recombinase" /note="FRT" CDS 2110..3132 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3262..3392 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3775..4448) /direction=LEFT /note="pUC ori" CDS complement(4593..5453) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5454..5552) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca aaaccgcggc tcttgcgcaa cacgatgaag ccgtggacaa caaattcaac 1141 aaagaacaac aaaacgcgtt ctatgagatc ttacatttac ctaacttaaa cgaagaacaa 1201 cgaaacgcct tcatccaaag tttaaaagat gacccaagcc aaagcgctaa ccttttagca 1261 gaagctaaaa agctaaatga tgctcaggcg ccgaaagtag acaacaaatt caacaaagaa 1321 caacaaaacg cgttctatga gatcttacat ttacctaact taaacgaaga acaacgaaac 1381 gccttcatcc aaagtttaaa agatgaccca agccaaagcg ctaacctttt agcagaagct 1441 aaaaagctaa atggtgctca ggcgccgaaa gtagacgcga attccgcggg gaagtcaacc 1501 tgactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt 1561 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact 1621 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat 1681 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc 1741 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc 1801 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt 1861 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc 1921 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct 1981 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat 2041 ggttcacgta cctagaagtt cctattccga agttcctatt ctctagaaag tataggaact 2101 tccttggcca aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag 2161 ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc 2221 ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac 2281 aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt 2341 gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc 2401 acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc 2461 atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg 2521 caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat 2581 gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc 2641 gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat 2701 ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc 2761 gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg 2821 ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga 2881 tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg 2941 gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga 3001 tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc 3061 gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg 3121 gcaaaggaat agcacgtact acgagatttc gattccaccg ccgccttcta tgaaaggttg 3181 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg 3241 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc 3301 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg 3361 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg 3421 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac 3481 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc 3541 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg 3601 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct 3661 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac 3721 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga 3781 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat 3841 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac 3901 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct 3961 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg 4021 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg 4081 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt 4141 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg 4201 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac 4261 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga 4321 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt 4381 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt 4441 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga 4501 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc 4561 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct 4621 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata 4681 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca 4741 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga 4801 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga 4861 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg 4921 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga 4981 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt 5041 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct 5101 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca 5161 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat 5221 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga 5281 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc 5341 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg 5401 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc 5461 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt 5521 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca 5581 cctgacgtc // >pKK-TEV-EGFP LOCUS Exported 5895 bp ds-DNA circular SYN 13-OCT-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Thursday, October 13, 2016 from SnapGene 3.2.1 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1809 /codon_start=1 /product="enhanced GFP" /note="EGFP" /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 1853..2077 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2361..2408 /bound_moiety="FLP recombinase" /note="FRT" CDS 2416..3438 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3568..3698 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4081..4754) /direction=LEFT /note="pUC ori" CDS complement(4899..5759) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5760..5858) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 1201 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 1381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag 1561 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg 1621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 1681 gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga gaagcgcgat 1741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg 1801 tacaagtaac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct 1861 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt 1921 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg 1981 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac 2041 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc 2101 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg 2161 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct 2221 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg 2281 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag 2341 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata 2401 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 2461 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 2521 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 2581 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa 2641 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 2701 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 2761 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc 2821 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 2881 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 2941 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 3001 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 3061 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 3121 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 3181 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 3241 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 3301 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc 3361 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 3421 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa 3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 5881 gtgccacctg acgtc // >pKK-TEV-mCherry LOCUS Exported 5886 bp ds-DNA circular SYN 14-PAŹ-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mCherry SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5886) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported 14 paza 2016 from SnapGene 3.0.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5886 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1800 /codon_start=1 /product="mCherry" /note="mCherry" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" polyA_signal 1844..2068 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2352..2399 /bound_moiety="FLP recombinase" /note="FRT" CDS 2407..3429 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3559..3689 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4072..4745) /direction=LEFT /note="pUC ori" CDS complement(4890..5750) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5751..5849) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg 1141 cgcttcaagg tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag 1201 ggcgagggcc gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc 1261 cccctgccct tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac 1321 gtgaagcacc ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag 1381 tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc 1441 ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac 1501 ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc 1561 gaggacggcg ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac 1621 tacgacgctg aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc 1681 tacaacgtca acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa 1741 cagtacgaac gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtag 1801 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc 1861 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc 1921 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct 1981 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg 2041 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct 2101 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg 2161 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct 2221 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta 2281 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt 2341 tcacgtacct agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc 2401 ttggccaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc 2461 gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc 2521 gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa 2581 gatcgttatg tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac 2641 attggggaat tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg 2701 ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg 2761 gatgcgatcg ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa 2821 ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg 2881 tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat 2941 gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc 3001 ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag 3061 gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg 3121 gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg 3181 ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt 3241 gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc 3301 ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat 3361 ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca 3421 aaggaatagc acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc 3481 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg 3541 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat 3601 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc 3661 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg 3721 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac 3781 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca 3841 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat 3901 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc 3961 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca 4021 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca 4081 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg 4141 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg 4201 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt 4261 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt 4321 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc 4381 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt 4441 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt 4501 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc 4561 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa 4621 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt 4681 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct 4741 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta 4801 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa 4861 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc 4921 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact 4981 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc 5041 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt 5101 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta 5161 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg 5221 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt 5281 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc 5341 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt 5401 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc 5461 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc 5521 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa 5581 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac 5641 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa 5701 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt 5761 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa 5821 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct 5881 gacgtc // >pKK-TEV-mCerulean LOCUS Exported 5895 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mCerulean SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1809 /codon_start=1 /product="Cerulean" /note="mCerulean" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 1853..2077 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2361..2408 /bound_moiety="FLP recombinase" /note="FRT" CDS 2416..3438 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3568..3698 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4081..4754) /direction=LEFT /note="pUC ori" CDS complement(4899..5759) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5760..5858) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 1201 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cctgacctgg ggcgtgcagt gcttcgcccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 1381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caacgccatc agcgacaacg tctatatcac cgccgacaag 1561 cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga cggcagcgtg 1621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 1681 gacaaccact acctgagcac ccagtccaag ctgagcaaag accccaacga gaagcgcgat 1741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg 1801 tacaagtaac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct 1861 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt 1921 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg 1981 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac 2041 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc 2101 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg 2161 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct 2221 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg 2281 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag 2341 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata 2401 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 2461 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 2521 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 2581 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa 2641 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 2701 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 2761 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc 2821 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 2881 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 2941 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 3001 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 3061 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 3121 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 3181 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 3241 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 3301 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc 3361 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 3421 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa 3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 5881 gtgccacctg acgtc // >pKK-TEV-BirA(R118G)-FLAG LOCUS Exported 6168 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-BirA(R118G)-FLAG SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6168) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..6168 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..2052 /organism="Escherichia coli str. K-12 substr. MG1655" /strain="K-12" /sub_strain="MG1655" /mol_type="genomic DNA" /db_xref="taxon:511145" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..2052 /codon_start=1 /transl_table=11 /gene="birA" /gene_synonym="bioR; dhbB; ECK3965; JW3941" /locus_tag="b3973" /product="bifunctional biotin-[acetylCoA carboxylase] holoenzyme synthetase/ DNA-binding transcriptional repressor, bio-5'-AMP-binding" /function="enzyme; Biosynthesis of cofactors, carriers: Biotin" /EC_number="6.3.4.15" /note="BirA(R118G)" /note="biotin-[acetylCoA carboxylase] holoenzyme synthetase and biotin operon repressor" /db_xref="GI:16131807" /db_xref="ASAP:ABE-0013001" /db_xref="UniProtKB/Swiss-Prot:P06709" /db_xref="EcoGene:EG10123" /db_xref="GeneID:948469" /protein_id="NP_418404.1" /translation="MKDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRD WGVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSG DACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRK LGADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQG WITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIG DKEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK" gene 1090..2052 /gene="birA" /gene_synonym="bioR; dhbB; ECK3965; JW3941" /locus_tag="b3973" /note="birA" /db_xref="EcoGene:EG10123" /db_xref="GeneID:948469" gene 1090..1093 /gene="murB" /gene_synonym="ECK3964; JW3940; yijB" /locus_tag="b3972" /note="murB" /db_xref="EcoGene:EG11205" /db_xref="GeneID:948470" CDS 2056..2082 /codon_start=1 /note="FLAG" /translation="DYKDDDDK" polyA_signal 2126..2350 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2634..2681 /bound_moiety="FLP recombinase" /note="FRT" CDS 2689..3711 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3841..3971 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4354..5027) /direction=LEFT /note="pUC ori" CDS complement(5172..6032) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(6033..6131) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tgaaggataa caccgtgcca ctgaaattga ttgccctgtt agcgaacggt 1141 gaatttcact ctggcgagca gttgggtgaa acgctgggaa tgagccgggc ggctattaat 1201 aaacacattc agacactgcg tgactggggc gttgatgtct ttaccgttcc gggtaaagga 1261 tacagcctgc ctgagcctat ccagttactt aatgctaaac agatattggg tcagctggat 1321 ggcggtagtg tagccgtgct gccagtgatt gactccacga atcagtacct tcttgatcgt 1381 atcggagagc ttaaatcggg cgatgcttgc attgcagaat accagcaggc tggccgtggt 1441 cgccggggtc ggaaatggtt ttcgcctttt ggcgcaaact tatatttgtc gatgttctgg 1501 cgtctggaac aaggcccggc ggcggcgatt ggtttaagtc tggttatcgg tatcgtgatg 1561 gcggaagtat tacgcaagct gggtgcagat aaagttcgtg ttaaatggcc taatgacctc 1621 tatctgcagg atcgcaagct ggcaggcatt ctggtggagc tgactggcaa aactggcgat 1681 gcggcgcaaa tagtcattgg agccgggatc aacatggcaa tgcgccgtgt tgaagagagt 1741 gtcgttaatc aggggtggat cacgctgcag gaagcgggga tcaatctcga tcgtaatacg 1801 ttggcggcca tgctaatacg tgaattacgt gctgcgttgg aactcttcga acaagaagga 1861 ttggcacctt atctgtcgcg ctgggaaaag ctggataatt ttattaatcg cccagtgaaa 1921 cttatcattg gtgataaaga aatatttggc atttcacgcg gaatagacaa acagggggct 1981 ttattacttg agcaggatgg aataataaaa ccctggatgg gcggtgaaat atccctgcgt 2041 agtgcagaaa aaatcgatta caaggatgac gacgataagt agctcgagtc tagagggccc 2101 gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc 2161 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa 2221 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg 2281 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg 2341 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg 2401 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca 2461 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc 2521 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct 2581 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc 2641 ctattccgaa gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa 2701 ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg 2761 atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga 2821 tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg 2881 cactttgcat cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag 2941 agcctgacct attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa 3001 accgaactgc ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc 3061 gatcttagcc agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact 3121 acatggcgtg atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg 3181 atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc 3241 gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg 3301 acggacaatg gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc 3361 caatacgagg tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag 3421 acgcgctact tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat 3481 atgctccgca ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat 3541 gcagcttggg cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg 3601 cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc 3661 gccgatagtg gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtacta 3721 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg 3781 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc 3841 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca 3901 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct 3961 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg 4021 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata 4081 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca 4141 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc 4201 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg 4261 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta 4321 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc 4381 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag 4441 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac 4501 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc 4561 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt 4621 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc 4681 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga 4741 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta 4801 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta 4861 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga 4921 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg 4981 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag 5041 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc 5101 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact 5161 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt 5221 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta 5281 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta 5341 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc 5401 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat 5461 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt 5521 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg 5581 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca 5641 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta 5701 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg 5761 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact 5821 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg 5881 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt 5941 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga 6001 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc 6061 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa 6121 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc // >pKK-TEV-mAmber LOCUS Exported 5895 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mAmber SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..1806 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1809 /codon_start=1 /note="mAmber" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGCGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 1853..2077 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2361..2408 /bound_moiety="FLP recombinase" /note="FRT" CDS 2416..3438 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3568..3698 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4081..4754) /direction=LEFT /note="pUC ori" CDS complement(4899..5759) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5760..5858) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 1201 gccacctacg gcaagctgac cctgaagctc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cctcggctgc ggcctgcagt gcttcgcccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 1381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caactacaac agccacaacg tctatatcac cgccgacaag 1561 cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga cggcggcgtg 1621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 1681 gacaaccact acctgagcta ccagtccaag ctgagcaaag accccaacga gaagcgcgat 1741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg 1801 tacaagtaac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct 1861 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt 1921 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg 1981 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac 2041 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc 2101 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg 2161 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct 2221 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg 2281 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag 2341 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata 2401 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 2461 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 2521 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 2581 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa 2641 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 2701 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 2761 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc 2821 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 2881 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 2941 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 3001 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 3061 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 3121 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 3181 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 3241 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 3301 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc 3361 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 3421 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa 3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 5881 gtgccacctg acgtc // >pKK-TEV-CyOFP1 LOCUS Exported 5880 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-CyOFP1 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5880) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5880 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1794 /codon_start=1 /note="CyOFP1" /translation="MVSKGEELIKENMRSKLYLEGSVNGHQFKCTHEGEGKPYEGKQTN RIKVVEGGPLPFAFDILATHFMYGSKVFIKYPADLPDYFKQSFPEGFTWERVMVFEDGG VLTATQDTSLQDGELIYNVKVRGVNFPANGPVMQKKTLGWEPSTETMYPADGGLEGRCD KALKLVGGGHLHVNFKTTYKSKKPVKMPGVHYVDRRLERIKEADNETYVEQYEHAVARY SNLGGGMDELYK" polyA_signal 1838..2062 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2346..2393 /bound_moiety="FLP recombinase" /note="FRT" CDS 2401..3423 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3553..3683 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4066..4739) /direction=LEFT /note="pUC ori" CDS complement(4884..5744) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5745..5843) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgatcaagg agaacatgag aagcaagctg 1141 tacctggaag gcagcgtgaa cggccaccag ttcaagtgca cccacgaagg ggagggcaag 1201 ccctacgagg gcaagcagac caacaggatc aaggtggtgg agggaggccc cctgccgttc 1261 gcattcgaca tcctggccac ccactttatg tacgggagca aggtgttcat caagtacccc 1321 gccgacctcc ccgattattt taagcagtcc ttccctgagg gcttcacatg ggagagagtc 1381 atggtgttcg aagacggggg cgtgctgacc gccacccagg acaccagcct ccaggacggc 1441 gagctcatct acaacgtcaa ggtcagaggg gtgaacttcc cagccaacgg ccccgtgatg 1501 cagaagaaaa cactgggctg ggagcccagc accgagacca tgtaccccgc tgacggcggc 1561 ctggaaggca gatgcgacaa ggccctgaag ctcgtgggcg ggggccacct gcacgtcaac 1621 ttcaagacca catacaagtc caagaaaccc gtgaagatgc ccggcgtcca ctacgtggac 1681 cgcagactgg aaagaatcaa ggaggccgac aacgagacct acgtcgagca gtacgagcac 1741 gctgtggcca gatactccaa cctgggcgga ggcatggacg agctgtacaa gtaactcgag 1801 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca 1861 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc 1921 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg 1981 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct 2041 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg 2101 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc 2161 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt 2221 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc 2281 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt 2341 acctagaagt tcctattccg aagttcctat tctctagaaa gtataggaac ttccttggcc 2401 aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc 2461 gtctccgacc tgatgcagct ctcggagggc gaagaatctc gtgctttcag cttcgatgta 2521 ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg atggtttcta caaagatcgt 2581 tatgtttatc ggcactttgc atcggccgcg ctcccgattc cggaagtgct tgacattggg 2641 gaattcagcg agagcctgac ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa 2701 gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg 2761 atcgctgcgg ccgatcttag ccagacgagc gggttcggcc cattcggacc gcaaggaatc 2821 ggtcaataca ctacatggcg tgatttcata tgcgcgattg ctgatcccca tgtgtatcac 2881 tggcaaactg tgatggacga caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg 2941 atgctttggg ccgaggactg ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc 3001 aacaatgtcc tgacggacaa tggccgcata acagcggtca ttgactggag cgaggcgatg 3061 ttcggggatt cccaatacga ggtcgccaac atcttcttct ggaggccgtg gttggcttgt 3121 atggagcagc agacgcgcta cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg 3181 ctccgggcgt atatgctccg cattggtctt gaccaactct atcagagctt ggttgacggc 3241 aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg caatcgtccg atccggagcc 3301 gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt 3361 gtagaagtac tcgccgatag tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa 3421 tagcacgtac tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga 3481 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc 3541 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc 3601 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc 3661 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca 3721 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga 3781 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt 3841 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga 3901 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc 3961 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg 4021 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc 4081 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc 4141 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga 4201 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc 4261 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat 4321 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg 4381 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc 4441 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga 4501 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact 4561 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt 4621 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag 4681 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg 4741 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa 4801 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata 4861 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg 4921 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata 4981 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg 5041 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct 5101 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt 5161 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc 5221 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga 5281 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt 5341 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc 5401 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa 5461 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca 5521 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca 5581 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct 5641 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc 5701 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa 5761 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt 5821 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc // >pKK-TEV-Dendra2N LOCUS Exported 5868 bp ds-DNA circular SYN 26-OCT-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-Dendra2N SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5868) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Monday, May 22, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5868 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..1779 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1779 /codon_start=1 /product="monomeric photoswitchable green-to-red fluorescent protein from Dendronephthya" /note="Dendra2" /note="mammalian codon-optimized" /translation="MNTPGINLIKEDMRVKVHMEGNVNGHAFVIEGEGKGKPYEGTQTA NLTVKEGAPLPFSYDILTTAVHYGNRVFTKYPEDIPDYFKQSFPEGYSWERTMTFEDKG ICTIRSDISLEGDCFFQNVRFKGTNFPPNGPVMQKKTLKWEPSTEKLHVRDGLLVGNIN MALLLEGGGHYLCDFKTTYKAKKVVQLPDAHFVDHRIEILGNDSDYNKVKLYEHAVARY SPLPSQVW" polyA_signal 1826..2050 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2334..2381 /bound_moiety="FLP recombinase" /note="FRT" CDS 2389..3411 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3541..3671 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4054..4727) /direction=LEFT /note="pUC ori" CDS complement(4872..5732) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5733..5831) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tgaacacccc gggaattaac ctgatcaagg aggacatgcg cgtgaaggtg 1141 cacatggagg gcaacgtgaa cggccacgcc ttcgtgatcg agggcgaggg caagggcaag 1201 ccctacgagg gcacccagac cgccaacctg accgtgaagg agggcgcccc cctgcccttc 1261 agctacgaca tcctgaccac cgccgtgcac tacggcaacc gggtgttcac caagtacccc 1321 gaggacatcc ccgactactt caagcagagc ttccccgagg gctacagctg ggagcgcacc 1381 atgaccttcg aggacaaggg catctgcacc atccgcagcg acatcagcct ggagggcgac 1441 tgcttcttcc agaacgtgcg cttcaagggc accaacttcc cccccaacgg ccccgtgatg 1501 cagaagaaga ccctgaagtg ggagcccagc accgagaagc tgcacgtgcg cgacggcctg 1561 ctggtgggca acatcaacat ggccctgctg ctggagggcg gcggccacta cctgtgcgac 1621 ttcaagacca cctacaaggc caagaaggtg gtgcagctgc ccgacgccca cttcgtggac 1681 caccgcatcg agatcctggg caacgacagc gactacaaca aggtgaagct gtacgagcac 1741 gccgtggccc gctacagccc cctgcccagc caggtgtggt aactcgagtc tagagggccc 1801 gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc 1861 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa 1921 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg 1981 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg 2041 ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta tccccacgcg 2101 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca 2161 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc 2221 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct 2281 ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtac ctagaagttc 2341 ctattccgaa gttcctattc tctagaaagt ataggaactt ccttggccaa aaagcctgaa 2401 ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt ctccgacctg 2461 atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg agggcgtgga 2521 tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta tgtttatcgg 2581 cactttgcat cggccgcgct cccgattccg gaagtgcttg acattgggga attcagcgag 2641 agcctgacct attgcatctc ccgccgtgca cagggtgtca cgttgcaaga cctgcctgaa 2701 accgaactgc ccgctgttct gcagccggtc gcggaggcca tggatgcgat cgctgcggcc 2761 gatcttagcc agacgagcgg gttcggccca ttcggaccgc aaggaatcgg tcaatacact 2821 acatggcgtg atttcatatg cgcgattgct gatccccatg tgtatcactg gcaaactgtg 2881 atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat gctttgggcc 2941 gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa caatgtcctg 3001 acggacaatg gccgcataac agcggtcatt gactggagcg aggcgatgtt cggggattcc 3061 caatacgagg tcgccaacat cttcttctgg aggccgtggt tggcttgtat ggagcagcag 3121 acgcgctact tcgagcggag gcatccggag cttgcaggat cgccgcggct ccgggcgtat 3181 atgctccgca ttggtcttga ccaactctat cagagcttgg ttgacggcaa tttcgatgat 3241 gcagcttggg cgcagggtcg atgcgacgca atcgtccgat ccggagccgg gactgtcggg 3301 cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt agaagtactc 3361 gccgatagtg gaaaccgacg ccccagcact cgtccgaggg caaaggaata gcacgtacta 3421 cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg 3481 gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccacccc 3541 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca 3601 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct 3661 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg 3721 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata 3781 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca 3841 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc 3901 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg 3961 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta 4021 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc 4081 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag 4141 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac 4201 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc 4261 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt 4321 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc 4381 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga 4441 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta 4501 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta 4561 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga 4621 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg 4681 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag 4741 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc 4801 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact 4861 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt 4921 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta 4981 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta 5041 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc 5101 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat 5161 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt 5221 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg 5281 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca 5341 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta 5401 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg 5461 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact 5521 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg 5581 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt 5641 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga 5701 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc 5761 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa 5821 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc // >pKK-TEV-mCardinal LOCUS Exported 5910 bp ds-DNA circular SYN 24-JAN-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mCardinal SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5910) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Monday, May 22, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5910 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..1821 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1821 /codon_start=1 /product="far-red fluorescent protein derived from mNeptune, with further red-shifted spectra" /note="mCardinal" /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTTEGEGKPYEGTQTQ RIKVVEGGPLPFAFDILATCFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG VLTVTQDTSLQDGCLIYNVKLRGVNFPSNGPVMQKKTLGWEATTETLYPADGGLEGRCD MALKLVGGGHLHCNLKTTYRSKKPAKNLKMPGVYFVDRRLERIKEADNETYVEQHEVAV ARYCDLPSKLGHKLNGMDELYK" polyA_signal 1868..2092 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2376..2423 /bound_moiety="FLP recombinase" /note="FRT" CDS 2431..3453 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3583..3713 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4096..4769) /direction=LEFT /note="pUC ori" CDS complement(4914..5774) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5775..5873) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgatcaagg agaacatgca catgaagctg 1141 tacatggaag gcaccgtgaa caaccaccac ttcaagtgca ccaccgaagg ggagggcaag 1201 ccctacgagg gcacccagac ccagaggatt aaggtggtgg agggaggccc cctgccgttc 1261 gcattcgaca tcctggccac ctgctttatg tacgggagca agaccttcat caaccacacc 1321 cagggcatcc ccgatttctt taagcagtcc ttccctgagg gcttcacatg ggagagagtc 1381 accacatacg aagacggggg cgtgcttacc gttacccagg acaccagcct ccaggacggc 1441 tgcttgatct acaacgtcaa gctcagaggg gtgaacttcc catccaacgg ccctgtgatg 1501 cagaagaaaa cactcggctg ggaggccacc accgagaccc tgtaccccgc tgacggcggc 1561 ctggaaggca gatgcgacat ggccctgaag ctcgtgggcg ggggccacct gcactgcaac 1621 ctgaagacca catacagatc caagaaaccc gctaagaacc tcaagatgcc cggcgtctac 1681 tttgtggacc gcagactgga aagaatcaag gaggccgaca atgagaccta cgtcgagcag 1741 cacgaggtgg ctgtggccag atactgcgac ctccctagca aactggggca caaacttaat 1801 ggcatggacg agctgtacaa gtaactcgag tctagagggc ccgtttaaac ccgctgatca 1861 gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc 1921 ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg 1981 cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg 2041 gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag 2101 gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta 2161 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg 2221 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa 2281 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc 2341 aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat 2401 tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc 2461 gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc 2521 gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat 2581 agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg 2641 ctcccgattc cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc 2701 tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt 2761 ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc 2821 gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata 2881 tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt 2941 gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc 3001 cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata 3061 acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac 3121 atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg 3181 aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt 3241 gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt 3301 cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc 3361 agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga 3421 cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc 3481 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc 3541 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct 3601 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca 3661 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg 3721 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt 3781 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt 3841 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg 3901 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg 3961 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg 4021 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat 4081 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 4141 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 4201 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga 4261 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt 4321 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg 4381 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc 4441 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg 4501 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc 4561 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg 4621 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc 4681 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct 4741 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt 4801 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa 4861 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa 4921 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc 4981 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct 5041 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca 5101 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt 5161 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt 5221 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc 5281 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc 5341 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt 5401 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact 5461 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc 5521 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt 5581 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg 5641 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct 5701 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa 5761 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt 5821 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc 5881 acatttcccc gaaaagtgcc acctgacgtc // >pKK-TEV-mClover3 LOCUS Exported 5895 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mClover3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1809 /codon_start=1 /note="mClover3" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" polyA_signal 1853..2077 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2361..2408 /bound_moiety="FLP recombinase" /note="FRT" CDS 2416..3438 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3568..3698 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4081..4754) /direction=LEFT /note="pUC ori" CDS complement(4899..5759) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5760..5858) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtcc gcggcgaggg cgagggcgat 1201 gccaccaacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cttcggctac ggcgtggcct gcttcagccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 1381 accatctctt tcaaggacga cggtacctac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caacttcaac agccactacg tctatatcac ggccgacaag 1561 cagaagaact gcatcaaggc taacttcaag atccgccaca acgttgagga cggcagcgtg 1621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 1681 gacaaccact acctgagcca tcagtccaag ctgagcaaag accccaacga gaagcgcgat 1741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatta cacatggcat ggacgagctg 1801 tacaagtaac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct 1861 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt 1921 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg 1981 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac 2041 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc 2101 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg 2161 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct 2221 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg 2281 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag 2341 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata 2401 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 2461 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 2521 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 2581 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa 2641 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 2701 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 2761 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc 2821 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 2881 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 2941 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 3001 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 3061 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 3121 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 3181 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 3241 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 3301 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc 3361 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 3421 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa 3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 5881 gtgccacctg acgtc // >pKK-TEV-mEGFP LOCUS Exported 5895 bp ds-DNA circular SYN 09-MAR-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mEGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Monday, May 22, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1809 /codon_start=1 /product="enhanced GFP" /note="mEGFP" /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 1853..2077 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2361..2408 /bound_moiety="FLP recombinase" /note="FRT" CDS 2416..3438 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3568..3698 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4081..4754) /direction=LEFT /note="pUC ori" CDS complement(4899..5759) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5760..5858) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 1201 gccacctacg gcaagctgac cctgaagttc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 1381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caactacaac agccacaacg tctatatcat ggccgacaag 1561 cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga cggcagcgtg 1621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 1681 gacaaccact acctgagcac ccagtccaag ctgagcaaag accccaacga gaagcgcgat 1741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg 1801 tacaagtaac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct 1861 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt 1921 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg 1981 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac 2041 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc 2101 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg 2161 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct 2221 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg 2281 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag 2341 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata 2401 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 2461 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 2521 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 2581 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa 2641 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 2701 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 2761 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc 2821 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 2881 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 2941 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 3001 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 3061 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 3121 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 3181 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 3241 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 3301 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc 3361 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 3421 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa 3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 5881 gtgccacctg acgtc // >pKK-TEV-mRuby3 LOCUS Exported 5889 bp ds-DNA circular SYN 24-JAN-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mRuby3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5889) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Monday, May 22, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5889 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1803 /codon_start=1 /note="mRuby3" /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV AKYSNLGGGMDELYK" polyA_signal 1847..2071 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2355..2402 /bound_moiety="FLP recombinase" /note="FRT" CDS 2410..3432 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3562..3692 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4075..4748) /direction=LEFT /note="pUC ori" CDS complement(4893..5753) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5754..5852) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgtctaa gggcgaagag ctgatcaagg aaaatatgcg tatgaaggtg 1141 gtcatggaag gttcggtcaa cggccaccaa ttcaaatgca caggtgaagg agaaggcaga 1201 ccgtacgagg gagtgcaaac catgaggatc aaagtcatcg agggaggacc cctgccattt 1261 gcctttgaca ttcttgccac gtcgttcatg tatggcagcc gtacctttat caagtacccg 1321 gccgacatcc ctgatttctt taaacagtcc tttcctgagg gttttacttg ggaaagagtt 1381 acgagatacg aagatggtgg agtcgtcacc gtcacgcagg acaccagcct tgaggatggc 1441 gagctcgtct acaacgtcaa ggtcagaggg gtaaactttc cctccaatgg tcccgtgatg 1501 cagaagaaga ccaagggttg ggagcctaat acagagatga tgtatccagc agatggtggt 1561 ctgagaggat acactgacat cgcactgaaa gttgatggtg gtggccatct gcactgcaac 1621 ttcgtgacaa cttacaggtc aaaaaagacc gtcgggaaca tcaagatgcc cggtgtccat 1681 gccgttgatc accgcctgga aaggatcgag gagagtgaca atgaaaccta cgtagtgcaa 1741 agagaagtgg cagttgccaa atacagcaac cttggtggtg gcatggacga gctgtacaag 1801 taactcgagt ctagagggcc cgtttaaacc cgctgatcag cctcgactgt gccttctagt 1861 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact 1921 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat 1981 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc 2041 aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac cagctggggc 2101 tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt 2161 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc 2221 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct 2281 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat 2341 ggttcacgta cctagaagtt cctattccga agttcctatt ctctagaaag tataggaact 2401 tccttggcca aaaagcctga actcaccgcg acgtctgtcg agaagtttct gatcgaaaag 2461 ttcgacagcg tctccgacct gatgcagctc tcggagggcg aagaatctcg tgctttcagc 2521 ttcgatgtag gagggcgtgg atatgtcctg cgggtaaata gctgcgccga tggtttctac 2581 aaagatcgtt atgtttatcg gcactttgca tcggccgcgc tcccgattcc ggaagtgctt 2641 gacattgggg aattcagcga gagcctgacc tattgcatct cccgccgtgc acagggtgtc 2701 acgttgcaag acctgcctga aaccgaactg cccgctgttc tgcagccggt cgcggaggcc 2761 atggatgcga tcgctgcggc cgatcttagc cagacgagcg ggttcggccc attcggaccg 2821 caaggaatcg gtcaatacac tacatggcgt gatttcatat gcgcgattgc tgatccccat 2881 gtgtatcact ggcaaactgt gatggacgac accgtcagtg cgtccgtcgc gcaggctctc 2941 gatgagctga tgctttgggc cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat 3001 ttcggctcca acaatgtcct gacggacaat ggccgcataa cagcggtcat tgactggagc 3061 gaggcgatgt tcggggattc ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg 3121 ttggcttgta tggagcagca gacgcgctac ttcgagcgga ggcatccgga gcttgcagga 3181 tcgccgcggc tccgggcgta tatgctccgc attggtcttg accaactcta tcagagcttg 3241 gttgacggca atttcgatga tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga 3301 tccggagccg ggactgtcgg gcgtacacaa atcgcccgca gaagcgcggc cgtctggacc 3361 gatggctgtg tagaagtact cgccgatagt ggaaaccgac gccccagcac tcgtccgagg 3421 gcaaaggaat agcacgtact acgagatttc gattccaccg ccgccttcta tgaaaggttg 3481 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg 3541 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc 3601 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg 3661 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg 3721 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac 3781 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc 3841 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg 3901 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct 3961 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac 4021 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga 4081 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat 4141 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac 4201 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct 4261 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg 4321 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg 4381 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt 4441 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg 4501 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac 4561 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga 4621 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt 4681 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt 4741 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga 4801 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc 4861 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct 4921 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata 4981 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca 5041 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga 5101 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga 5161 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg 5221 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga 5281 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt 5341 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct 5401 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca 5461 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat 5521 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga 5581 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc 5641 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg 5701 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc 5761 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt 5821 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca 5881 cctgacgtc // >pKK-TEV-mTagBFP LOCUS Exported 5877 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mTagBFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5877) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5877 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..1788 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1788 /codon_start=1 /product="monomeric blue fluorescent protein" /note="mTagBFP" /note="mammalian codon-optimized" /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC DLPSKLGHKLN" polyA_signal 1835..2059 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2343..2390 /bound_moiety="FLP recombinase" /note="FRT" CDS 2398..3420 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3550..3680 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4063..4736) /direction=LEFT /note="pUC ori" CDS complement(4881..5741) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5742..5840) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tgagcgagct gattaaggag aacatgcaca tgaagctgta catggagggc 1141 accgtggaca accatcactt caagtgcaca tccgagggcg aaggcaagcc ctacgagggc 1201 acccagacca tgagaatcaa ggtggtcgag ggcggccctc tccccttcgc cttcgacatc 1261 ctggctacta gcttcctcta cggcagcaag accttcatca accacaccca gggcatcccc 1321 gacttcttca agcagtcctt ccctgagggc ttcacatggg agagagtcac cacatacgaa 1381 gacgggggcg tgctgaccgc tacccaggac accagcctcc aggacggctg cctcatctac 1441 aacgtcaaga tcagaggggt gaacttcaca tccaacggcc ctgtgatgca gaagaaaaca 1501 ctcggctggg aggccttcac cgagacgctg taccccgctg acggcggcct ggaaggcaga 1561 aacgacatgg ccctgaagct cgtgggcggg agccatctga tcgcaaacat caagaccaca 1621 tatagatcca agaaacccgc taagaacctc aagatgcctg gcgtctacta tgtggactac 1681 agactggaaa gaatcaagga ggccaacaac gagacctacg tcgagcagca cgaggtggca 1741 gtggccagat actgcgacct ccctagcaaa ctggggcaca agcttaatta actcgagtct 1801 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct 1861 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt 1921 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg 1981 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg 2041 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat 2101 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg 2161 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc 2221 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga 2281 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtacc 2341 tagaagttcc tattccgaag ttcctattct ctagaaagta taggaacttc cttggccaaa 2401 aagcctgaac tcaccgcgac gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc 2461 tccgacctga tgcagctctc ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga 2521 gggcgtggat atgtcctgcg ggtaaatagc tgcgccgatg gtttctacaa agatcgttat 2581 gtttatcggc actttgcatc ggccgcgctc ccgattccgg aagtgcttga cattggggaa 2641 ttcagcgaga gcctgaccta ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac 2701 ctgcctgaaa ccgaactgcc cgctgttctg cagccggtcg cggaggccat ggatgcgatc 2761 gctgcggccg atcttagcca gacgagcggg ttcggcccat tcggaccgca aggaatcggt 2821 caatacacta catggcgtga tttcatatgc gcgattgctg atccccatgt gtatcactgg 2881 caaactgtga tggacgacac cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg 2941 ctttgggccg aggactgccc cgaagtccgg cacctcgtgc acgcggattt cggctccaac 3001 aatgtcctga cggacaatgg ccgcataaca gcggtcattg actggagcga ggcgatgttc 3061 ggggattccc aatacgaggt cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg 3121 gagcagcaga cgcgctactt cgagcggagg catccggagc ttgcaggatc gccgcggctc 3181 cgggcgtata tgctccgcat tggtcttgac caactctatc agagcttggt tgacggcaat 3241 ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg 3301 actgtcgggc gtacacaaat cgcccgcaga agcgcggccg tctggaccga tggctgtgta 3361 gaagtactcg ccgatagtgg aaaccgacgc cccagcactc gtccgagggc aaaggaatag 3421 cacgtactac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc 3481 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc 3541 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca 3601 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc 3661 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg 3721 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc 3781 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg 3841 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc 3901 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact 3961 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta 4021 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag 4081 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc 4141 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta 4201 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg 4261 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc 4321 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac 4381 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac 4441 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg 4501 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga 4561 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt 4621 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag 4681 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct 4741 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg 4801 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat 4861 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc 4921 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg 4981 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct 5041 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca 5101 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg 5161 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg 5221 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc 5281 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag 5341 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg 5401 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag 5461 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat 5521 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg 5581 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca 5641 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca 5701 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat 5761 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag 5821 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc // >pKK-TEV-mVenus LOCUS Exported 5895 bp ds-DNA circular SYN 26-OCT-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-TEV-mVenus SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5895) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5895 /organism="synthetic DNA construct" /mol_type="other DNA" source 1090..1806 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1090..1806 /codon_start=1 /product="Venus YFP with monomerizing A206K mutation (Nagai et al., 2002)" /note="mVenus" /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 1853..2077 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2361..2408 /bound_moiety="FLP recombinase" /note="FRT" CDS 2416..3438 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3568..3698 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(4081..4754) /direction=LEFT /note="pUC ori" CDS complement(4899..5759) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5760..5858) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctggcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatca tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc catcctggtc 1141 gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg cgagggcgat 1201 gccacctacg gcaagctgac cctgaagctc atctgcacca ccggcaagct gcccgtgccc 1261 tggcccaccc tcgtgaccac cctcggctac ggcctgcagt gcttcgcccg ctaccccgac 1321 cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt ccaggagcgc 1381 accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa gttcgagggc 1441 gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga cggcaacatc 1501 ctggggcaca agctggagta caactacaac agccacaacg tctatatcac cgccgacaag 1561 cagaagaacg gcatcaaggc caacttcaag atccgccaca acatcgagga cggcggcgtg 1621 cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt gctgctgccc 1681 gacaaccact acctgagcta ccagtccaag ctgagcaaag accccaacga gaagcgcgat 1741 cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat ggacgagctg 1801 tacaagtaac tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct 1861 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt 1921 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg 1981 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac 2041 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc 2101 tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg 2161 gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct 2221 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg 2281 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag 2341 ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct agaaagtata 2401 ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 2461 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 2521 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 2581 ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc gattccggaa 2641 gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 2701 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 2761 gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt cggcccattc 2821 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 2881 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 2941 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 3001 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 3061 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 3121 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 3181 gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 3241 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 3301 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc 3361 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 3421 ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc cttctatgaa 3481 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat 3541 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa 3601 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt 3661 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag 3721 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt 3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc 3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc 3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct 3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca 4021 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac 4081 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt 4141 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg 4201 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc 4261 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc 4321 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc 4381 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac 4441 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt 4501 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct 4561 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc 4621 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt 4681 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg 4741 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc 4801 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa 4861 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag 4921 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg 4981 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga 5041 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag 5101 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa 5161 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc 5221 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca 5281 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg 5341 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat 5401 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc 5461 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg 5521 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg 5581 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt 5641 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca 5701 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata 5761 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac 5821 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa 5881 gtgccacctg acgtc // >pKK-BI16-TEV-mCherry LOCUS Exported 6060 bp ds-DNA circular SYN 24-OCT-2016 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-BI16-TEV-mCherry SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6060) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6060) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Wednesday, May 24, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6060 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" misc_feature 557..633 /note="MCS" misc_feature 636..1163 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 641..659 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 662..680 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1119..1137 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1140..1158 /bound_moiety="Tet repressor" /note="tet operator" CDS 1188..1208 /codon_start=1 /product="enocdes peptide cleaved by TEV protease" /note="TEV-L" /translation="ENLYFQG" misc_feature 1249..1269 /product="enocdes peptide cleaved by TEV protease" /note="TEV-R" CDS 1276..1986 /codon_start=1 /product="mCherry" /note="mCherry" /note="mCherry color: #ff0000" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" polyA_signal 2018..2242 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2526..2573 /bound_moiety="FLP recombinase" /note="FRT" CDS 2581..3603 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3733..3863 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4246..4919) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5064..5924) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(5925..6023) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc ccttatctac gcgtttcaga gcggccgcac tgatcttaag 601 ggtactagcg ctgatcggca attgcgaggt cctaggacga tctctatcac tgatagggag 661 atctctatca ctgataggga gagctctgct tatatagacc tcccaccgta cacgcctacc 721 gcccatttgc gtcaatgggg cggagttgtt acgacatttt ggaaagtccc gttgattttg 781 gtgccaaaac aaactcccat tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc 841 aaaccgctat ccacgcccat tgatgtactg ccaaaaccgc atcaccatgg taatagcgat 901 gactaatacg tagatgtact gccaagtagg aaagtcccat aaggtcatgt actgggcata 961 atgccaggcg ggccatttac cgtcattgac gtcaataggg ggcgtacttg gcatatgata 1021 cacttgatgt actctagaaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg 1081 taggcgtgta cggtgggagg tctatataag cagagctctc cctatcagtg atagagatct 1141 ccctatcagt gatagagatc gtcaagcttg gtaccgagct cggatccgaa aacctgtact 1201 tccaaggaac cggtggtcac ctgtacaatc gatcctgcag gagctagcga gaatttgtat 1261 tttcagggtg atatcatggt gagcaagggc gaggaggata acatggccat catcaaggag 1321 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag 1381 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag 1441 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag 1501 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc 1561 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac 1621 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc 1681 tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg 1741 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc 1801 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc 1861 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc 1921 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac 1981 aagtagctcg aggtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca 2041 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc 2101 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg 2161 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct 2221 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg 2281 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc 2341 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt 2401 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc 2461 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt 2521 acctagaagt tcctattccg aagttcctat tctctagaaa gtataggaac ttccttggcc 2581 aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc 2641 gtctccgacc tgatgcagct ctcggagggc gaagaatctc gtgctttcag cttcgatgta 2701 ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg atggtttcta caaagatcgt 2761 tatgtttatc ggcactttgc atcggccgcg ctcccgattc cggaagtgct tgacattggg 2821 gaattcagcg agagcctgac ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa 2881 gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg 2941 atcgctgcgg ccgatcttag ccagacgagc gggttcggcc cattcggacc gcaaggaatc 3001 ggtcaataca ctacatggcg tgatttcata tgcgcgattg ctgatcccca tgtgtatcac 3061 tggcaaactg tgatggacga caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg 3121 atgctttggg ccgaggactg ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc 3181 aacaatgtcc tgacggacaa tggccgcata acagcggtca ttgactggag cgaggcgatg 3241 ttcggggatt cccaatacga ggtcgccaac atcttcttct ggaggccgtg gttggcttgt 3301 atggagcagc agacgcgcta cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg 3361 ctccgggcgt atatgctccg cattggtctt gaccaactct atcagagctt ggttgacggc 3421 aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg caatcgtccg atccggagcc 3481 gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt 3541 gtagaagtac tcgccgatag tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa 3601 tagcacgtac tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga 3661 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc 3721 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc 3781 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc 3841 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca 3901 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga 3961 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt 4021 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga 4081 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc 4141 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg 4201 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc 4261 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc 4321 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga 4381 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc 4441 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat 4501 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg 4561 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc 4621 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga 4681 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact 4741 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt 4801 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag 4861 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg 4921 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa 4981 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata 5041 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg 5101 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata 5161 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg 5221 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct 5281 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt 5341 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc 5401 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga 5461 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt 5521 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc 5581 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa 5641 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca 5701 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca 5761 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct 5821 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc 5881 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa 5941 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt 6001 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc // >pKK-BI16-FLAG-C3-ORF1_mClover3-TEV-ORF2 LOCUS Exported 6126 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-BI16-FLAG-C3-ORF1_mClover3-TEV-ORF2 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6126) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6126) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6126 /organism="synthetic DNA construct" /mol_type="other DNA" source 1219..2052 /organism="synthetic DNA construct" /mol_type="other DNA" source 1225..1941 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" misc_feature 557..624 /note="MCS" CDS complement(625..648) /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /note="HRV 3C site" /translation="LEVLFQGP" CDS complement(649..672) /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /note="FLAG" /translation="DYKDDDDK" misc_feature 685..1212 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 690..708 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 711..729 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1168..1186 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1189..1207 /bound_moiety="Tet repressor" /note="tet operator" CDS 1225..1941 /codon_start=1 /note="mClover3" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" misc_feature 1948..1968 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 2009..2029 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 2084..2308 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2592..2639 /bound_moiety="FLP recombinase" /note="FRT" CDS 2647..3669 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3799..3929 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4312..4985) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5130..5990) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(5991..6089) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc ccttatctac gcgtttcaga gcggccgcac tgatcttaag 601 ggtactagcg ctgatcggca attggggccc ctggaacaga acttccagct tatcgtcgtc 661 atccttgtaa tccataggtc ctaggacgat ctctatcact gatagggaga tctctatcac 721 tgatagggag agctctgctt atatagacct cccaccgtac acgcctaccg cccatttgcg 781 tcaatggggc ggagttgtta cgacattttg gaaagtcccg ttgattttgg tgccaaaaca 841 aactcccatt gacgtcaatg gggtggagac ttggaaatcc ccgtgagtca aaccgctatc 901 cacgcccatt gatgtactgc caaaaccgca tcaccatggt aatagcgatg actaatacgt 961 agatgtactg ccaagtagga aagtcccata aggtcatgta ctgggcataa tgccaggcgg 1021 gccatttacc gtcattgacg tcaatagggg gcgtacttgg catatgatac acttgatgta 1081 ctctagaaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac 1141 ggtgggaggt ctatataagc agagctctcc ctatcagtga tagagatctc cctatcagtg 1201 atagagatcg tcaagcttgg taccatggtg agcaagggcg aggagctgtt caccggggtg 1261 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtccgcggc 1321 gagggcgagg gcgatgccac caacggcaag ctgaccctga agttcatctg caccaccggc 1381 aagctgcccg tgccctggcc caccctcgtg accaccttcg gctacggcgt ggcctgcttc 1441 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc 1501 tacgtccagg agcgcaccat ctctttcaag gacgacggta cctacaagac ccgcgccgag 1561 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag 1621 gaggacggca acatcctggg gcacaagctg gagtacaact tcaacagcca ctacgtctat 1681 atcacggccg acaagcagaa gaactgcatc aaggctaact tcaagatccg ccacaacgtt 1741 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc 1801 cccgtgctgc tgcccgacaa ccactacctg agccatcagt ccaagctgag caaagacccc 1861 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gattacacat 1921 ggcatggacg agctgtacaa gggatccgaa aacctgtact tccaaggaac cggtggtcac 1981 ctgtacaatc gatcctgcag gagctagcga gaatttgtat tttcagggtg atatccagca 2041 cagtggcggc cgctcgaggt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc 2101 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc 2161 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct 2221 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg 2281 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct 2341 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg 2401 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct 2461 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta 2521 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt 2581 tcacgtacct agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc 2641 ttggccaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc 2701 gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc 2761 gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa 2821 gatcgttatg tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac 2881 attggggaat tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg 2941 ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg 3001 gatgcgatcg ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa 3061 ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg 3121 tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat 3181 gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc 3241 ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag 3301 gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg 3361 gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg 3421 ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt 3481 gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc 3541 ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat 3601 ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca 3661 aaggaatagc acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc 3721 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg 3781 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat 3841 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc 3901 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg 3961 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac 4021 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca 4081 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat 4141 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc 4201 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca 4261 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca 4321 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg 4381 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg 4441 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt 4501 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt 4561 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc 4621 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt 4681 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt 4741 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc 4801 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa 4861 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt 4921 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct 4981 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta 5041 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa 5101 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc 5161 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact 5221 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc 5281 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt 5341 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta 5401 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg 5461 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt 5521 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc 5581 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt 5641 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc 5701 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc 5761 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa 5821 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac 5881 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa 5941 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt 6001 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa 6061 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct 6121 gacgtc // >pKK-BI16-ORF1-C3-mRuby3_ORF2-TEV-mClover3 LOCUS Exported 6800 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-BI16-ORF1-C3-mRuby3_ORF2-TEV-mClover3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6800) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6800) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6800 /organism="synthetic DNA construct" /mol_type="other DNA" source 562..1275 /organism="synthetic DNA construct" /mol_type="other DNA" source 1901..2726 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" CDS complement(562..1275) /codon_start=1 /note="mRuby3" /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV AKYSNLGGGMDELYK" CDS complement(1276..1299) /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /note="HRV 3C site" /translation="LEVLFQGP" misc_feature 1300..1364 /note="MCS" misc_feature 1367..1894 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 1372..1390 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1393..1411 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1850..1868 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1871..1889 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 1919..1939 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1980..2000 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 2007..2726 /codon_start=1 /note="mClover3" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" polyA_signal 2758..2982 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3266..3313 /bound_moiety="FLP recombinase" /note="FRT" CDS 3321..4343 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4473..4603 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4986..5659) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5804..6664) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6665..6763) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cttacttgta cagctcgtcc atgccaccac caaggttgct 601 gtatttggca actgccactt ctctttgcac tacgtaggtt tcattgtcac tctcctcgat 661 cctttccagg cggtgatcaa cggcatggac accgggcatc ttgatgttcc cgacggtctt 721 ttttgacctg taagttgtca cgaagttgca gtgcagatgg ccaccaccat caactttcag 781 tgcgatgtca gtgtatcctc tcagaccacc atctgctgga tacatcatct ctgtattagg 841 ctcccaaccc ttggtcttct tctgcatcac gggaccattg gagggaaagt ttacccctct 901 gaccttgacg ttgtagacga gctcgccatc ctcaaggctg gtgtcctgcg tgacggtgac 961 gactccacca tcttcgtatc tcgtaactct ttcccaagta aaaccctcag gaaaggactg 1021 tttaaagaaa tcagggatgt cggccgggta cttgataaag gtacggctgc catacatgaa 1081 cgacgtggca agaatgtcaa aggcaaatgg caggggtcct ccctcgatga ctttgatcct 1141 catggtttgc actccctcgt acggtctgcc ttctccttca cctgtgcatt tgaattggtg 1201 gccgttgacc gaaccttcca tgaccacctt catacgcata ttttccttga tcagctcttc 1261 gcccttagac accatgggcc cctggaacag aacttccaga cgcgtttcag agcggccgca 1321 ctgatcttaa gggtactagc gctgatcggc aattgcgagg tcctaggacg atctctatca 1381 ctgataggga gatctctatc actgataggg agagctctgc ttatatagac ctcccaccgt 1441 acacgcctac cgcccatttg cgtcaatggg gcggagttgt tacgacattt tggaaagtcc 1501 cgttgatttt ggtgccaaaa caaactccca ttgacgtcaa tggggtggag acttggaaat 1561 ccccgtgagt caaaccgcta tccacgccca ttgatgtact gccaaaaccg catcaccatg 1621 gtaatagcga tgactaatac gtagatgtac tgccaagtag gaaagtccca taaggtcatg 1681 tactgggcat aatgccaggc gggccattta ccgtcattga cgtcaatagg gggcgtactt 1741 ggcatatgat acacttgatg tactctagaa aaatgtcgta acaactccgc cccattgacg 1801 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt 1861 gatagagatc tccctatcag tgatagagat cgtcaagctt ggtaccgagc tcggatccga 1921 aaacctgtac ttccaaggaa ccggtggtca cctgtacaat cgatcctgca ggagctagcg 1981 agaatttgta ttttcagggt gatatcatgg tgagcaaggg cgaggagctg ttcaccgggg 2041 tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtccgcg 2101 gcgagggcga gggcgatgcc accaacggca agctgaccct gaagttcatc tgcaccaccg 2161 gcaagctgcc cgtgccctgg cccaccctcg tgaccacctt cggctacggc gtggcctgct 2221 tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag 2281 gctacgtcca ggagcgcacc atctctttca aggacgacgg tacctacaag acccgcgccg 2341 aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca 2401 aggaggacgg caacatcctg gggcacaagc tggagtacaa cttcaacagc cactacgtct 2461 atatcacggc cgacaagcag aagaactgca tcaaggctaa cttcaagatc cgccacaacg 2521 ttgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg 2581 gccccgtgct gctgcccgac aaccactacc tgagccatca gtccaagctg agcaaagacc 2641 ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggattacac 2701 atggcatgga cgagctgtac aagtaactcg aggtttaaac ccgctgatca gcctcgactg 2761 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg 2821 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga 2881 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg 2941 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa 3001 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg 3061 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt 3121 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc 3181 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg 3241 attagggtga tggttcacgt acctagaagt tcctattccg aagttcctat tctctagaaa 3301 gtataggaac ttccttggcc aaaaagcctg aactcaccgc gacgtctgtc gagaagtttc 3361 tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct ctcggagggc gaagaatctc 3421 gtgctttcag cttcgatgta ggagggcgtg gatatgtcct gcgggtaaat agctgcgccg 3481 atggtttcta caaagatcgt tatgtttatc ggcactttgc atcggccgcg ctcccgattc 3541 cggaagtgct tgacattggg gaattcagcg agagcctgac ctattgcatc tcccgccgtg 3601 cacagggtgt cacgttgcaa gacctgcctg aaaccgaact gcccgctgtt ctgcagccgg 3661 tcgcggaggc catggatgcg atcgctgcgg ccgatcttag ccagacgagc gggttcggcc 3721 cattcggacc gcaaggaatc ggtcaataca ctacatggcg tgatttcata tgcgcgattg 3781 ctgatcccca tgtgtatcac tggcaaactg tgatggacga caccgtcagt gcgtccgtcg 3841 cgcaggctct cgatgagctg atgctttggg ccgaggactg ccccgaagtc cggcacctcg 3901 tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa tggccgcata acagcggtca 3961 ttgactggag cgaggcgatg ttcggggatt cccaatacga ggtcgccaac atcttcttct 4021 ggaggccgtg gttggcttgt atggagcagc agacgcgcta cttcgagcgg aggcatccgg 4081 agcttgcagg atcgccgcgg ctccgggcgt atatgctccg cattggtctt gaccaactct 4141 atcagagctt ggttgacggc aatttcgatg atgcagcttg ggcgcagggt cgatgcgacg 4201 caatcgtccg atccggagcc gggactgtcg ggcgtacaca aatcgcccgc agaagcgcgg 4261 ccgtctggac cgatggctgt gtagaagtac tcgccgatag tggaaaccga cgccccagca 4321 ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt cgattccacc gccgccttct 4381 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg 4441 gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt 4501 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta 4561 gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta 4621 gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca 4681 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag 4741 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt 4801 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc 4861 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg 4921 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa 4981 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg 5041 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga 5101 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg 5161 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg 5221 gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc 5281 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg 5341 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca 5401 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt 5461 ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag 5521 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg 5581 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc 5641 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt 5701 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt 5761 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca 5821 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg 5881 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac 5941 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg 6001 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc 6061 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta 6121 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac 6181 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc 6241 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac 6301 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact 6361 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa 6421 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt 6481 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca 6541 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa 6601 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac 6661 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg 6721 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc 6781 gaaaagtgcc acctgacgtc // >pKK-BI16-ORF1-C3-FLAG_ORF2-TEV-mClover3 LOCUS Exported 6116 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-BI16-ORF1-C3-FLAG_ORF2-TEV-mClover3 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6116) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6116) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6116 /organism="synthetic DNA construct" /mol_type="other DNA" source 565..591 /organism="synthetic DNA construct" /mol_type="other DNA" source 1217..2042 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" CDS complement(565..591) /codon_start=1 /product="FLAG" /note="FLAG" /translation="MDYKDDDDK" CDS complement(592..615) /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /note="HRV 3C site" /translation="LEVLFQGP" misc_feature 616..680 /note="MCS" misc_feature 683..1210 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 688..706 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 709..727 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1166..1184 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1187..1205 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 1235..1255 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1296..1316 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 1323..2039 /codon_start=1 /note="mClover3" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTL KFICTTGKLPVPWPTLVTTFGYGVACFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDD GTYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHYVYITADKQKNCIK ANFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSHQSKLSKDPNEKRDHMVLL EFVTAAGITHGMDELYK" polyA_signal 2074..2298 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2582..2629 /bound_moiety="FLP recombinase" /note="FRT" CDS 2637..3659 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3789..3919 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4302..4975) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5120..5980) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(5981..6079) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cttacttatc gtcgtcatcc ttgtaatcca tgggcccctg 601 gaacagaact tccagacgcg tttcagagcg gccgcactga tcttaagggt actagcgctg 661 atcggcaatt gcgaggtcct aggacgatct ctatcactga tagggagatc tctatcactg 721 atagggagag ctctgcttat atagacctcc caccgtacac gcctaccgcc catttgcgtc 781 aatggggcgg agttgttacg acattttgga aagtcccgtt gattttggtg ccaaaacaaa 841 ctcccattga cgtcaatggg gtggagactt ggaaatcccc gtgagtcaaa ccgctatcca 901 cgcccattga tgtactgcca aaaccgcatc accatggtaa tagcgatgac taatacgtag 961 atgtactgcc aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc 1021 catttaccgt cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact 1081 ctagaaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg 1141 tgggaggtct atataagcag agctctccct atcagtgata gagatctccc tatcagtgat 1201 agagatcgtc aagcttggta ccgagctcgg atccgaaaac ctgtacttcc aaggaaccgg 1261 tggtcacctg tacaatcgat cctgcaggag ctagcgagaa tttgtatttt cagggtgata 1321 tcatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg 1381 acggcgacgt aaacggccac aagttcagcg tccgcggcga gggcgagggc gatgccacca 1441 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca 1501 ccctcgtgac caccttcggc tacggcgtgg cctgcttcag ccgctacccc gaccacatga 1561 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct 1621 ctttcaagga cgacggtacc tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc 1681 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc 1741 acaagctgga gtacaacttc aacagccact acgtctatat cacggccgac aagcagaaga 1801 actgcatcaa ggctaacttc aagatccgcc acaacgttga ggacggcagc gtgcagctcg 1861 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc 1921 actacctgag ccatcagtcc aagctgagca aagaccccaa cgagaagcgc gatcacatgg 1981 tcctgctgga gttcgtgacc gccgccggga ttacacatgg catggacgag ctgtacaagt 2041 aactcgaggt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg 2101 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt 2161 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg 2221 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg 2281 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc 2341 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 2401 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 2461 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 2521 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtacct 2581 agaagttcct attccgaagt tcctattctc tagaaagtat aggaacttcc ttggccaaaa 2641 agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct 2701 ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag 2761 ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg 2821 tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat 2881 tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc 2941 tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg 3001 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc 3061 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc 3121 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc 3181 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca 3241 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg 3301 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg 3361 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc 3421 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt 3481 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga 3541 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag 3601 aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca aaggaatagc 3661 acgtactacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg 3721 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg 3781 cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa 3841 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca 3901 atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt 3961 catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg 4021 gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt 4081 tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg 4141 gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg 4201 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa 4261 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc 4321 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc 4381 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat 4441 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc 4501 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct 4561 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg 4621 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc 4681 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga 4741 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa 4801 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta 4861 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc 4921 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg 4981 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga 5041 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg 5101 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct 5161 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg 5221 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 5281 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa 5341 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 5401 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 5461 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 5521 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt 5581 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 5641 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 5701 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 5761 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga 5821 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 5881 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 5941 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt 6001 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga 6061 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc // >pKK-BiFC-Venus LOCUS Exported 6141 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-BiFC-Venus SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6141) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6141) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6141 /organism="synthetic DNA construct" /mol_type="other DNA" source 557..562 /organism="synthetic DNA construct" /mol_type="other DNA" source 563..814 /organism="pBiFC-VC155 Sequence" /mol_type="other DNA" source 1546..2067 /organism="pBiFC-VN173 Sequence" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" CDS complement(563..814) /codon_start=1 /note="Venus 155-239" /translation="KQKNGIKANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHY LSYQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYK" CDS complement(815..838) /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /note="HRV 3C site" /translation="LEVLFQGP" misc_feature 839..903 /note="MCS" misc_feature 906..1433 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 911..929 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 932..950 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1389..1407 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1410..1428 /bound_moiety="Tet repressor" /note="tet operator" CDS 1458..1478 /codon_start=1 /product="enocdes peptide cleaved by TEV protease" /note="TEV-L" /translation="ENLYFQG" misc_feature 1519..1539 /product="enocdes peptide cleaved by TEV protease" /note="TEV-R" CDS 1546..2067 /codon_start=1 /note="Venus N1-173" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIE" polyA_signal 2099..2323 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 2607..2654 /bound_moiety="FLP recombinase" /note="FRT" CDS 2662..3684 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 3814..3944 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4327..5000) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5145..6005) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6006..6104) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc ccttacttgt acagctcgtc catgccgaga gtgatcccgg 601 cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca 661 gtttggactg gtagctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg 721 gggtgttctg ctggtagtgg tcggcgagct gcacgccgcc gtcctcgatg ttgtggcgga 781 tcttgaagtt ggccttgatg ccgttcttct gcttgggccc ctggaacaga acttccagac 841 gcgtttcaga gcggccgcac tgatcttaag ggtactagcg ctgatcggca attgcgaggt 901 cctaggacga tctctatcac tgatagggag atctctatca ctgataggga gagctctgct 961 tatatagacc tcccaccgta cacgcctacc gcccatttgc gtcaatgggg cggagttgtt 1021 acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat tgacgtcaat 1081 ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat tgatgtactg 1141 ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact gccaagtagg 1201 aaagtcccat aaggtcatgt actgggcata atgccaggcg ggccatttac cgtcattgac 1261 gtcaataggg ggcgtacttg gcatatgata cacttgatgt actctagaaa aatgtcgtaa 1321 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag 1381 cagagctctc cctatcagtg atagagatct ccctatcagt gatagagatc gtcaagcttg 1441 gtaccgagct cggatccgaa aacctgtact tccaaggaac cggtggtcac ctgtacaatc 1501 gatcctgcag gagctagcga gaatttgtat tttcagggtg atatcatggt gagcaagggc 1561 gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc 1621 cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa gctgaccctg 1681 aagctgatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt gaccaccctg 1741 ggctacggcc tgcagtgctt cgcccgctac cccgaccaca tgaagcagca cgacttcttc 1801 aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa ggacgacggc 1861 aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag 1921 ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct ggagtacaac 1981 tacaacagcc acaacgtcta tatcaccgcc gacaagcaga agaacggcat caaggccaac 2041 ttcaagatcc gccacaacat cgagtagctc gaggtttaaa cccgctgatc agcctcgact 2101 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg 2161 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg 2221 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg 2281 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga 2341 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg 2401 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct 2461 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat 2521 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt 2581 gattagggtg atggttcacg tacctagaag ttcctattcc gaagttccta ttctctagaa 2641 agtataggaa cttccttggc caaaaagcct gaactcaccg cgacgtctgt cgagaagttt 2701 ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct 2761 cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc 2821 gatggtttct acaaagatcg ttatgtttat cggcactttg catcggccgc gctcccgatt 2881 ccggaagtgc ttgacattgg ggaattcagc gagagcctga cctattgcat ctcccgccgt 2941 gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg 3001 gtcgcggagg ccatggatgc gatcgctgcg gccgatctta gccagacgag cgggttcggc 3061 ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt 3121 gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc 3181 gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc 3241 gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc 3301 attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc 3361 tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg 3421 gagcttgcag gatcgccgcg gctccgggcg tatatgctcc gcattggtct tgaccaactc 3481 tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac 3541 gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg 3601 gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc 3661 actcgtccga gggcaaagga atagcacgta ctacgagatt tcgattccac cgccgccttc 3721 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc 3781 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt 3841 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct 3901 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct 3961 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc 4021 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga 4081 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg 4141 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg 4201 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg 4261 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga 4321 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg 4381 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag 4441 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc 4501 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg 4561 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt 4621 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc 4681 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc 4741 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg 4801 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca 4861 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc 4921 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat 4981 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt 5041 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt 5101 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc 5161 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc 5221 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata 5281 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg 5341 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc 5401 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct 5461 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa 5521 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt 5581 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca 5641 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac 5701 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca 5761 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt 5821 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc 5881 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca 5941 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata 6001 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc 6061 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc 6121 cgaaaagtgc cacctgacgt c // >pKK-FRET-ORF1-C3-mCerulean_ORF2-TEV-mVenus LOCUS Exported 6807 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-FRET-Orf1-C3-mCerulean_Orf2-TEV-mVenus SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6807) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6807) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6807 /organism="synthetic DNA construct" /mol_type="other DNA" source 557..562 /organism="synthetic DNA construct" /mol_type="other DNA" source 563..1282 /organism="synthetic DNA construct" /mol_type="other DNA" source 1908..2733 /organism="synthetic DNA construct" /mol_type="other DNA" source 2014..2730 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" CDS complement(563..1282) /codon_start=1 /product="Cerulean" /note="mCerulean" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS complement(1283..1306) /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /note="HRV 3C site" /translation="LEVLFQGP" misc_feature 1307..1371 /note="MCS" misc_feature 1374..1901 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 1379..1397 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1400..1418 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1857..1875 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1878..1896 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 1926..1946 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1987..2007 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 2014..2733 /codon_start=1 /note="mVenus" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 2765..2989 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3273..3320 /bound_moiety="FLP recombinase" /note="FRT" CDS 3328..4350 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4480..4610 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4993..5666) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5811..6671) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6672..6770) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc ccttacttgt acagctcgtc catgccgaga gtgatcccgg 601 cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca 661 gcttggactg ggtgctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg 721 gggtgttctg ctggtagtgg tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga 781 tcttgaagtt ggccttgatg ccgttcttct gcttgtcggc ggtgatatag acgttgtcgc 841 tgatggcgtt gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc 901 ccttcagctc gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct 961 tgtagttgcc gtcgtccttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg 1021 cggacttgaa gaagtcgtgc tgcttcatgt ggtcggggta gcgggcgaag cactgcacgc 1081 cccaggtcag ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga 1141 tgaacttcag ggtcagcttg ccgtaggtgg catcgccctc gccctcgccg gacacgctga 1201 acttgtggcc gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca 1261 gctcctcgcc cttgctcacc atgggcccct ggaacagaac ttccagacgc gtttcagagc 1321 ggccgcactg atcttaaggg tactagcgct gatcggcaat tgcgaggtcc taggacgatc 1381 tctatcactg atagggagat ctctatcact gatagggaga gctctgctta tatagacctc 1441 ccaccgtaca cgcctaccgc ccatttgcgt caatggggcg gagttgttac gacattttgg 1501 aaagtcccgt tgattttggt gccaaaacaa actcccattg acgtcaatgg ggtggagact 1561 tggaaatccc cgtgagtcaa accgctatcc acgcccattg atgtactgcc aaaaccgcat 1621 caccatggta atagcgatga ctaatacgta gatgtactgc caagtaggaa agtcccataa 1681 ggtcatgtac tgggcataat gccaggcggg ccatttaccg tcattgacgt caataggggg 1741 cgtacttggc atatgataca cttgatgtac tctagaaaaa tgtcgtaaca actccgcccc 1801 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc 1861 tatcagtgat agagatctcc ctatcagtga tagagatcgt caagcttggt accgagctcg 1921 gatccgaaaa cctgtacttc caaggaaccg gtggtcacct gtacaatcga tcctgcagga 1981 gctagcgaga atttgtattt tcagggtgat atcatggtga gcaagggcga ggagctgttc 2041 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc 2101 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gctcatctgc 2161 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctcgg ctacggcctg 2221 cagtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg 2281 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc 2341 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc 2401 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac 2461 aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc 2521 cacaacatcg aggacggcgg cgtgcagctc gccgaccact accagcagaa cacccccatc 2581 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gctaccagtc caagctgagc 2641 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg 2701 atcactctcg gcatggacga gctgtacaag taactcgagg tttaaacccg ctgatcagcc 2761 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg 2821 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat 2881 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag 2941 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg 3001 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc 3061 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc 3121 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct 3181 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa 3241 aaacttgatt agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct 3301 ctagaaagta taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag 3361 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa 3421 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc 3481 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc 3541 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc 3601 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg 3661 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg 3721 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc 3781 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg 3841 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg 3901 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca 3961 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc 4021 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg 4081 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac 4141 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga 4201 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga 4261 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc 4321 cccagcactc gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc 4381 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc 4441 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat 4501 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg 4561 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg 4621 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat 4681 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc 4741 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga 4801 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt 4861 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg 4921 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac 4981 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg 5041 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca 5101 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc 5161 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc 5221 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag 5281 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc 5341 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca 5401 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg 5461 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg 5521 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct 5581 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa 5641 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa 5701 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa 5761 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc 5821 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga 5881 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca 5941 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc 6001 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat 6061 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc 6121 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt 6181 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc 6241 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg 6301 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt 6361 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg 6421 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga 6481 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg 6541 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg 6601 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt 6661 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc 6721 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca 6781 tttccccgaa aagtgccacc tgacgtc // >pKK-FRET-ORF1-C3-mCerulean_ORF2-TEV-mAmber LOCUS Exported 6807 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-FRET-ORF1-C3-mCerulean_ORF2-TEV-mAmber SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6807) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6807) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 ORIGDB|GenBank FEATURES Location/Qualifiers source 1..6807 /organism="synthetic DNA construct" /mol_type="other DNA" source 557..562 /organism="synthetic DNA construct" /mol_type="other DNA" source 563..1282 /organism="synthetic DNA construct" /mol_type="other DNA" source 1908..2733 /organism="synthetic DNA construct" /mol_type="other DNA" source 2014..2730 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" /note="TK polyadenylation signal" CDS complement(563..1282) /codon_start=1 /product="Cerulean" /note="Cerulean" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAISDNVYITADKQKNGIK ANFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" CDS complement(1283..1306) /codon_start=1 /product="recognition and cleavage site for human rhinovirus 3C and PreScission proteases" /note="HRV 3C site" /translation="LEVLFQGP" misc_feature 1307..1371 /note="MCS" misc_feature 1374..1901 /note="BI Promoter" /note="bidirectional promoter from BI16 vector (Sammarco and Grabczyk, PMID: 16212928)" protein_bind 1379..1397 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1400..1418 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1857..1875 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1878..1896 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 1926..1946 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1987..2007 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" CDS 2014..2733 /codon_start=1 /note="mAmber" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KLICTTGKLPVPWPTLVTTLGCGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSKLSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 2765..2989 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3273..3320 /bound_moiety="FLP recombinase" /note="FRT" CDS 3328..4350 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4480..4610 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4993..5666) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5811..6671) /codon_start=1 /product="beta-lactamase" /note="AmpR" /note="Ampicillin color: #ff9966" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6672..6770) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc ccttacttgt acagctcgtc catgccgaga gtgatcccgg 601 cggcggtcac gaactccagc aggaccatgt gatcgcgctt ctcgttgggg tctttgctca 661 gcttggactg ggtgctcagg tagtggttgt cgggcagcag cacggggccg tcgccgatgg 721 gggtgttctg ctggtagtgg tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga 781 tcttgaagtt ggccttgatg ccgttcttct gcttgtcggc ggtgatatag acgttgtcgc 841 tgatggcgtt gtactccagc ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc 901 ccttcagctc gatgcggttc accagggtgt cgccctcgaa cttcacctcg gcgcgggtct 961 tgtagttgcc gtcgtccttg aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg 1021 cggacttgaa gaagtcgtgc tgcttcatgt ggtcggggta gcgggcgaag cactgcacgc 1081 cccaggtcag ggtggtcacg agggtgggcc agggcacggg cagcttgccg gtggtgcaga 1141 tgaacttcag ggtcagcttg ccgtaggtgg catcgccctc gccctcgccg gacacgctga 1201 acttgtggcc gtttacgtcg ccgtccagct cgaccaggat gggcaccacc ccggtgaaca 1261 gctcctcgcc cttgctcacc atgggcccct ggaacagaac ttccagacgc gtttcagagc 1321 ggccgcactg atcttaaggg tactagcgct gatcggcaat tgcgaggtcc taggacgatc 1381 tctatcactg atagggagat ctctatcact gatagggaga gctctgctta tatagacctc 1441 ccaccgtaca cgcctaccgc ccatttgcgt caatggggcg gagttgttac gacattttgg 1501 aaagtcccgt tgattttggt gccaaaacaa actcccattg acgtcaatgg ggtggagact 1561 tggaaatccc cgtgagtcaa accgctatcc acgcccattg atgtactgcc aaaaccgcat 1621 caccatggta atagcgatga ctaatacgta gatgtactgc caagtaggaa agtcccataa 1681 ggtcatgtac tgggcataat gccaggcggg ccatttaccg tcattgacgt caataggggg 1741 cgtacttggc atatgataca cttgatgtac tctagaaaaa tgtcgtaaca actccgcccc 1801 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc 1861 tatcagtgat agagatctcc ctatcagtga tagagatcgt caagcttggt accgagctcg 1921 gatccgaaaa cctgtacttc caaggaaccg gtggtcacct gtacaatcga tcctgcagga 1981 gctagcgaga atttgtattt tcagggtgat atcatggtga gcaagggcga ggagctgttc 2041 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc 2101 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gctcatctgc 2161 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctcgg ctgcggcctg 2221 cagtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg 2281 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc 2341 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc 2401 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac 2461 aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc 2521 cacaacatcg aggacggcgg cgtgcagctc gccgaccact accagcagaa cacccccatc 2581 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gctaccagtc caagctgagc 2641 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg 2701 atcactctcg gcatggacga gctgtacaag taactcgagg tttaaacccg ctgatcagcc 2761 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg 2821 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat 2881 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag 2941 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg 3001 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc 3061 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc 3121 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct 3181 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa 3241 aaacttgatt agggtgatgg ttcacgtacc tagaagttcc tattccgaag ttcctattct 3301 ctagaaagta taggaacttc cttggccaaa aagcctgaac tcaccgcgac gtctgtcgag 3361 aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc ggagggcgaa 3421 gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg ggtaaatagc 3481 tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc ggccgcgctc 3541 ccgattccgg aagtgcttga cattggggaa ttcagcgaga gcctgaccta ttgcatctcc 3601 cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc cgctgttctg 3661 cagccggtcg cggaggccat ggatgcgatc gctgcggccg atcttagcca gacgagcggg 3721 ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga tttcatatgc 3781 gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac cgtcagtgcg 3841 tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc cgaagtccgg 3901 cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg ccgcataaca 3961 gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt cgccaacatc 4021 ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt cgagcggagg 4081 catccggagc ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac 4141 caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga 4201 tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga 4261 agcgcggccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc 4321 cccagcactc gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc 4381 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc 4441 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat 4501 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg 4561 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg 4621 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat 4681 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc 4741 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga 4801 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt 4861 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg 4921 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac 4981 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg 5041 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca 5101 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc 5161 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc 5221 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag 5281 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc 5341 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca 5401 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg 5461 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg 5521 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct 5581 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa 5641 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa 5701 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa 5761 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc 5821 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga 5881 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca 5941 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc 6001 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat 6061 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc 6121 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt 6181 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc 6241 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg 6301 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt 6361 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg 6421 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga 6481 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg 6541 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg 6601 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt 6661 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc 6721 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca 6781 tttccccgaa aagtgccacc tgacgtc // >pKK-RNAi-nucEGFPmiR-FLAG-TEV LOCUS Exported 6618 bp ds-DNA circular SYN 24-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucEGFPmiR-FLAG-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6618) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6618) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..6618 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1138..1852) /codon_start=1 /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELY" misc_feature 1861..2388 /note="BI Promoter" protein_bind 1866..1884 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1887..1905 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1969..1984 /note="100F" primer_bind 2240..2258 /note="372F" primer_bind 2265..2280 /note="100F" protein_bind 2344..2362 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2365..2383 /bound_moiety="Tet repressor" /note="tet operator" CDS 2407..2433 /codon_start=1 /product="FLAG" /note="FLAG" /translation="MDYKDDDDK" misc_feature 2440..2460 /label=TEV /note="TEV-L" misc_feature 2501..2521 /label=TEV /note="TEV-R" polyA_signal 2576..2800 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3084..3131 /bound_moiety="FLP recombinase" /note="FRT" CDS 3139..4161 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4291..4421 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4804..5477) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5622..6482) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6483..6581) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt 1201 gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt 1261 cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct 1321 gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct 1381 gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca 1441 ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt 1501 cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg 1561 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt 1621 ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc 1681 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg 1741 catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct 1801 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtcctag 1861 gacgatctct atcactgata gggagatctc tatcactgat agggagagct ctgcttatat 1921 agacctccca ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag ttgttacgac 1981 attttggaaa gtcccgttga ttttggtgcc aaaacaaact cccattgacg tcaatggggt 2041 ggagacttgg aaatccccgt gagtcaaacc gctatccacg cccattgatg tactgccaaa 2101 accgcatcac catggtaata gcgatgacta atacgtagat gtactgccaa gtaggaaagt 2161 cccataaggt catgtactgg gcataatgcc aggcgggcca tttaccgtca ttgacgtcaa 2221 tagggggcgt acttggcata tgatacactt gatgtactct agaaaaatgt cgtaacaact 2281 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag 2341 ctctccctat cagtgataga gatctcccta tcagtgatag agatcgtcgg atctaagctt 2401 ggtaccatgg attacaagga tgacgacgat aagggatccg aaaacctgta cttccaagga 2461 accggtggtc acctgtacaa tcgatcctgc aggagctagc gagaatttgt attttcaggg 2521 tgatatccag cacagtggca gctgctcgag gtttaaaccc gctgatcagc ctcgactgtg 2581 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa 2641 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt 2701 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa 2761 gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc 2821 agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt 2881 gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc 2941 gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg 3001 gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat 3061 tagggtgatg gttcacgtac ctagaagttc ctattccgaa gttcctattc tctagaaagt 3121 ataggaactt ccttggccaa aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg 3181 atcgaaaagt tcgacagcgt ctccgacctg atgcagctct cggagggcga agaatctcgt 3241 gctttcagct tcgatgtagg agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat 3301 ggtttctaca aagatcgtta tgtttatcgg cactttgcat cggccgcgct cccgattccg 3361 gaagtgcttg acattgggga attcagcgag agcctgacct attgcatctc ccgccgtgca 3421 cagggtgtca cgttgcaaga cctgcctgaa accgaactgc ccgctgttct gcagccggtc 3481 gcggaggcca tggatgcgat cgctgcggcc gatcttagcc agacgagcgg gttcggccca 3541 ttcggaccgc aaggaatcgg tcaatacact acatggcgtg atttcatatg cgcgattgct 3601 gatccccatg tgtatcactg gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg 3661 caggctctcg atgagctgat gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg 3721 cacgcggatt tcggctccaa caatgtcctg acggacaatg gccgcataac agcggtcatt 3781 gactggagcg aggcgatgtt cggggattcc caatacgagg tcgccaacat cttcttctgg 3841 aggccgtggt tggcttgtat ggagcagcag acgcgctact tcgagcggag gcatccggag 3901 cttgcaggat cgccgcggct ccgggcgtat atgctccgca ttggtcttga ccaactctat 3961 cagagcttgg ttgacggcaa tttcgatgat gcagcttggg cgcagggtcg atgcgacgca 4021 atcgtccgat ccggagccgg gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc 4081 gtctggaccg atggctgtgt agaagtactc gccgatagtg gaaaccgacg ccccagcact 4141 cgtccgaggg caaaggaata gcacgtacta cgagatttcg attccaccgc cgccttctat 4201 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg 4261 gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta taatggttac 4321 aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact gcattctagt 4381 tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc gacctctagc 4441 tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca 4501 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg 4561 agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg 4621 tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc 4681 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta 4741 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag 4801 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg 4861 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg 4921 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg 4981 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga 5041 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc 5101 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt 5161 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact 5221 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg 5281 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt 5341 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt 5401 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct 5461 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg 5521 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt 5581 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt 5641 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc 5701 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg 5761 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc 5821 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg 5881 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca 5941 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga 6001 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct 6061 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg 6121 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca 6181 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata 6241 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct 6301 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact 6361 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa 6421 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc 6481 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga 6541 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga 6601 aaagtgccac ctgacgtc // >pKK-RNAi-nucEGFPmiR-mCherry-TEV LOCUS Exported 7293 bp ds-DNA circular SYN 26-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucEGFPmiR-mCherry-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7293) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 7293) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..7293 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1138..1852) /codon_start=1 /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELY" misc_feature 1861..2388 /note="BI Promoter" protein_bind 1866..1884 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1887..1905 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1969..1984 /note="100F" primer_bind 2240..2258 /note="372F" primer_bind 2265..2280 /note="100F" protein_bind 2344..2362 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2365..2383 /bound_moiety="Tet repressor" /note="tet operator" CDS 2401..3108 /codon_start=1 /label=mCherry /note="mCherry" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" misc_feature 3115..3135 /label=TEV /note="TEV-L" misc_feature 3176..3196 /label=TEV /note="TEV-R" polyA_signal 3251..3475 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3759..3806 /bound_moiety="FLP recombinase" /note="FRT" CDS 3814..4836 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4966..5096 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(5479..6152) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(6297..7157) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(7158..7256) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt 1201 gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt 1261 cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct 1321 gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct 1381 gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca 1441 ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt 1501 cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg 1561 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt 1621 ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc 1681 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg 1741 catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct 1801 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtcctag 1861 gacgatctct atcactgata gggagatctc tatcactgat agggagagct ctgcttatat 1921 agacctccca ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag ttgttacgac 1981 attttggaaa gtcccgttga ttttggtgcc aaaacaaact cccattgacg tcaatggggt 2041 ggagacttgg aaatccccgt gagtcaaacc gctatccacg cccattgatg tactgccaaa 2101 accgcatcac catggtaata gcgatgacta atacgtagat gtactgccaa gtaggaaagt 2161 cccataaggt catgtactgg gcataatgcc aggcgggcca tttaccgtca ttgacgtcaa 2221 tagggggcgt acttggcata tgatacactt gatgtactct agaaaaatgt cgtaacaact 2281 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag 2341 ctctccctat cagtgataga gatctcccta tcagtgatag agatcgtcaa gcttggtacc 2401 atggtgagca agggcgagga ggataacatg gccatcatca aggagttcat gcgcttcaag 2461 gtgcacatgg agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc 2521 cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc 2581 ttcgcctggg acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac 2641 cccgccgaca tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc 2701 gtgatgaact tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac 2761 ggcgagttca tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta 2821 atgcagaaga agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc 2881 gccctgaagg gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgct 2941 gaggtcaaga ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc 3001 aacatcaagt tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa 3061 cgcgccgagg gccgccactc caccggcggc atggacgagc tgtacaaggg atccgaaaac 3121 ctgtacttcc aaggaaccgg tggtcacctg tacaatcgat cctgcaggag ctagcgagaa 3181 tttgtatttt cagggtgata tccagcacag tggcagctgc tcgaggttta aacccgctga 3241 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct 3301 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca 3361 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag 3421 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct 3481 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca 3541 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta 3601 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt 3661 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac 3721 cccaaaaaac ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc 3781 tattctctag aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct 3841 gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag 3901 ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta 3961 aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc 4021 gcgctcccga ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc 4081 atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct 4141 gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg 4201 agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc 4261 atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc 4321 agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa 4381 gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc 4441 ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc 4501 aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag 4561 cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt 4621 cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag 4681 ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc 4741 cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac 4801 cgacgcccca gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc 4861 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg 4921 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca 4981 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt 5041 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata 5101 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat 5161 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg 5221 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag 5281 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt 5341 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg 5401 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg 5461 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag 5521 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga 5581 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct 5641 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc 5701 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg 5761 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc 5821 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca 5881 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag 5941 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct 6001 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc 6061 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga 6121 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca 6181 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat 6241 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac 6301 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt 6361 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt 6421 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag 6481 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct 6541 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt 6601 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc 6661 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt 6721 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg 6781 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg 6841 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct 6901 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc 6961 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt 7021 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt 7081 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg 7141 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat 7201 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg 7261 cgcacatttc cccgaaaagt gccacctgac gtc // >pKK-RNAi-nucCHERRYmiR-FLAG-TEV LOCUS Exported 6603 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucCHERRY miR-FLAG-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6603) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6603) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..6603 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1144..1852) /codon_start=1 /note="mCherry" /translation="PRTMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRP YEGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERV MNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDG ALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQY ERAEGRHSTGGMDE" misc_feature 1852..2379 /note="BI Promoter" protein_bind 1857..1875 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1878..1896 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1960..1975 /note="100F" primer_bind 2231..2249 /note="372F" primer_bind 2256..2271 /note="100F" protein_bind 2335..2353 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2356..2374 /bound_moiety="Tet repressor" /note="tet operator" CDS 2392..2418 /codon_start=1 /product="FLAG" /note="FLAG" /translation="MDYKDDDDK" misc_feature 2425..2445 /label=TEV /note="TEV-L" misc_feature 2486..2506 /label=TEV /note="TEV-R" polyA_signal 2561..2785 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3069..3116 /bound_moiety="FLP recombinase" /note="FRT" CDS 3124..4146 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4276..4406 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4789..5462) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5607..6467) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6468..6566) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgccg gtggagtggc ggccctcggc gcgttcgtac tgttccacga 1201 tggtgtagtc ctcgttgtgg gaggtgatgt ccaacttgat gttgacgttg taggcgccgg 1261 gcagctgcac gggcttcttg gccttgtagg tggtcttgac ctcagcgtcg tagtggccgc 1321 cgtccttcag cttcagcctc tgcttgatct cgcccttcag ggcgccgtcc tcggggtaca 1381 tccgctcgga ggaggcctcc cagcccatgg tcttcttctg cattacgggg ccgtcggagg 1441 ggaagttggt gccgcgcagc ttcaccttgt agatgaactc gccgtcctgc agggaggagt 1501 cctgggtcac ggtcaccacg ccgccgtcct cgaagttcat cacgcgctcc cacttgaagc 1561 cctcggggaa ggacagcttc aagtagtcgg ggatgtcggc ggggtgcttc acgtaggcct 1621 tggagccgta catgaactga ggggacagga tgtcccaggc gaagggcagg gggccaccct 1681 tggtcacctt cagcttggcg gtctgggtgc cctcgtaggg gcggccctcg ccctcgccct 1741 cgatctcgaa ctcgtggccg ttcacggagc cctccatgtg caccttgaag cgcatgaact 1801 ccttgatgat ggccatgtta tcctcctcgc ccttgctcac catggtccta ggacgatctc 1861 tatcactgat agggagatct ctatcactga tagggagagc tctgcttata tagacctccc 1921 accgtacacg cctaccgccc atttgcgtca atggggcgga gttgttacga cattttggaa 1981 agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg tggagacttg 2041 gaaatccccg tgagtcaaac cgctatccac gcccattgat gtactgccaa aaccgcatca 2101 ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag tcccataagg 2161 tcatgtactg ggcataatgc caggcgggcc atttaccgtc attgacgtca atagggggcg 2221 tacttggcat atgatacact tgatgtactc tagaaaaatg tcgtaacaac tccgccccat 2281 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctcccta 2341 tcagtgatag agatctccct atcagtgata gagatcgtca agcttggtac catggattac 2401 aaggatgacg acgataaggg atccgaaaac ctgtacttcc aaggaaccgg tggtcacctg 2461 tacaatcgat cctgcaggag ctagcgagaa tttgtatttt cagggtgata tccagcacag 2521 tggcagctgc tcgaggttta aacccgctga tcagcctcga ctgtgccttc tagttgccag 2581 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact 2641 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt 2701 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat 2761 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg 2821 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc 2881 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc 2941 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg 3001 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca 3061 cgtacctaga agttcctatt ccgaagttcc tattctctag aaagtatagg aacttccttg 3121 gccaaaaagc ctgaactcac cgcgacgtct gtcgagaagt ttctgatcga aaagttcgac 3181 agcgtctccg acctgatgca gctctcggag ggcgaagaat ctcgtgcttt cagcttcgat 3241 gtaggagggc gtggatatgt cctgcgggta aatagctgcg ccgatggttt ctacaaagat 3301 cgttatgttt atcggcactt tgcatcggcc gcgctcccga ttccggaagt gcttgacatt 3361 ggggaattca gcgagagcct gacctattgc atctcccgcc gtgcacaggg tgtcacgttg 3421 caagacctgc ctgaaaccga actgcccgct gttctgcagc cggtcgcgga ggccatggat 3481 gcgatcgctg cggccgatct tagccagacg agcgggttcg gcccattcgg accgcaagga 3541 atcggtcaat acactacatg gcgtgatttc atatgcgcga ttgctgatcc ccatgtgtat 3601 cactggcaaa ctgtgatgga cgacaccgtc agtgcgtccg tcgcgcaggc tctcgatgag 3661 ctgatgcttt gggccgagga ctgccccgaa gtccggcacc tcgtgcacgc ggatttcggc 3721 tccaacaatg tcctgacgga caatggccgc ataacagcgg tcattgactg gagcgaggcg 3781 atgttcgggg attcccaata cgaggtcgcc aacatcttct tctggaggcc gtggttggct 3841 tgtatggagc agcagacgcg ctacttcgag cggaggcatc cggagcttgc aggatcgccg 3901 cggctccggg cgtatatgct ccgcattggt cttgaccaac tctatcagag cttggttgac 3961 ggcaatttcg atgatgcagc ttgggcgcag ggtcgatgcg acgcaatcgt ccgatccgga 4021 gccgggactg tcgggcgtac acaaatcgcc cgcagaagcg cggccgtctg gaccgatggc 4081 tgtgtagaag tactcgccga tagtggaaac cgacgcccca gcactcgtcc gagggcaaag 4141 gaatagcacg tactacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc 4201 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag 4261 ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc 4321 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa 4381 ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa 4441 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata 4501 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta 4561 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa 4621 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg 4681 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag 4741 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa 4801 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc 4861 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca 4921 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg 4981 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct 5041 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt 5101 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag 5161 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc 5221 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac 5281 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga 5341 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc 5401 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg 5461 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca 5521 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt 5581 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca 5641 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg 5701 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca 5761 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt 5821 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt 5881 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca 5941 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca 6001 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga 6061 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact 6121 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga 6181 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg 6241 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc 6301 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga 6361 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat 6421 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt 6481 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt 6541 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac 6601 gtc // >pKK-RNAi-nucCHERRYmiR-EGFP-TEV LOCUS Exported 7293 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucCHERRYmiR-EGFP-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7293) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 7293) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..7293 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1144..1852) /codon_start=1 /note="mCherry" /translation="PRTMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRP YEGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERV MNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDG ALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQY ERAEGRHSTGGMDE" misc_feature 1852..2379 /note="BI Promoter" protein_bind 1857..1875 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1878..1896 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1960..1975 /note="100F" primer_bind 2231..2249 /note="372F" primer_bind 2256..2271 /note="100F" protein_bind 2335..2353 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2356..2374 /bound_moiety="Tet repressor" /note="tet operator" CDS 2392..3108 /codon_start=1 /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" misc_feature 3115..3135 /label=TEV /note="TEV-L" misc_feature 3176..3196 /label=TEV /note="TEV-R" polyA_signal 3251..3475 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3759..3806 /bound_moiety="FLP recombinase" /note="FRT" CDS 3814..4836 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4966..5096 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(5479..6152) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(6297..7157) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(7158..7256) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgccg gtggagtggc ggccctcggc gcgttcgtac tgttccacga 1201 tggtgtagtc ctcgttgtgg gaggtgatgt ccaacttgat gttgacgttg taggcgccgg 1261 gcagctgcac gggcttcttg gccttgtagg tggtcttgac ctcagcgtcg tagtggccgc 1321 cgtccttcag cttcagcctc tgcttgatct cgcccttcag ggcgccgtcc tcggggtaca 1381 tccgctcgga ggaggcctcc cagcccatgg tcttcttctg cattacgggg ccgtcggagg 1441 ggaagttggt gccgcgcagc ttcaccttgt agatgaactc gccgtcctgc agggaggagt 1501 cctgggtcac ggtcaccacg ccgccgtcct cgaagttcat cacgcgctcc cacttgaagc 1561 cctcggggaa ggacagcttc aagtagtcgg ggatgtcggc ggggtgcttc acgtaggcct 1621 tggagccgta catgaactga ggggacagga tgtcccaggc gaagggcagg gggccaccct 1681 tggtcacctt cagcttggcg gtctgggtgc cctcgtaggg gcggccctcg ccctcgccct 1741 cgatctcgaa ctcgtggccg ttcacggagc cctccatgtg caccttgaag cgcatgaact 1801 ccttgatgat ggccatgtta tcctcctcgc ccttgctcac catggtccta ggacgatctc 1861 tatcactgat agggagatct ctatcactga tagggagagc tctgcttata tagacctccc 1921 accgtacacg cctaccgccc atttgcgtca atggggcgga gttgttacga cattttggaa 1981 agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg tggagacttg 2041 gaaatccccg tgagtcaaac cgctatccac gcccattgat gtactgccaa aaccgcatca 2101 ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag tcccataagg 2161 tcatgtactg ggcataatgc caggcgggcc atttaccgtc attgacgtca atagggggcg 2221 tacttggcat atgatacact tgatgtactc tagaaaaatg tcgtaacaac tccgccccat 2281 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctcccta 2341 tcagtgatag agatctccct atcagtgata gagatcgtca agcttggtac catggtgagc 2401 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta 2461 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg 2521 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc 2581 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac 2641 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac 2701 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc 2761 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag 2821 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag 2881 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac 2941 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc 3001 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag 3061 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggg atccgaaaac 3121 ctgtacttcc aaggaaccgg tggtcacctg tacaatcgat cctgcaggag ctagcgagaa 3181 tttgtatttt cagggtgata tccagcacag tggcagctgc tcgaggttta aacccgctga 3241 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct 3301 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca 3361 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag 3421 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct 3481 gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca 3541 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta 3601 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt 3661 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac 3721 cccaaaaaac ttgattaggg tgatggttca cgtacctaga agttcctatt ccgaagttcc 3781 tattctctag aaagtatagg aacttccttg gccaaaaagc ctgaactcac cgcgacgtct 3841 gtcgagaagt ttctgatcga aaagttcgac agcgtctccg acctgatgca gctctcggag 3901 ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta 3961 aatagctgcg ccgatggttt ctacaaagat cgttatgttt atcggcactt tgcatcggcc 4021 gcgctcccga ttccggaagt gcttgacatt ggggaattca gcgagagcct gacctattgc 4081 atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct 4141 gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg 4201 agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacatg gcgtgatttc 4261 atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc 4321 agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa 4381 gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc 4441 ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc 4501 aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag 4561 cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt 4621 cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag 4681 ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc 4741 cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac 4801 cgacgcccca gcactcgtcc gagggcaaag gaatagcacg tactacgaga tttcgattcc 4861 accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg 4921 atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca 4981 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt 5041 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata 5101 ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat 5161 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg 5221 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag 5281 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt 5341 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg 5401 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg 5461 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag 5521 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga 5581 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct 5641 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc 5701 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg 5761 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc 5821 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca 5881 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag 5941 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct 6001 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc 6061 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga 6121 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca 6181 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat 6241 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac 6301 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt 6361 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt 6421 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag 6481 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct 6541 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt 6601 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc 6661 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt 6721 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg 6781 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg 6841 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct 6901 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc 6961 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt 7021 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt 7081 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg 7141 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat 7201 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg 7261 cgcacatttc cccgaaaagt gccacctgac gtc // >pKK-RNAi-nucEGFPmiR-TEV-FLAG LOCUS Exported 6601 bp ds-DNA circular SYN 26-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucEGFPmiR-TEV-FLAG SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6601) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6601) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..6601 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1138..1852) /codon_start=1 /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELY" misc_feature 1861..2388 /note="BI Promoter" protein_bind 1866..1884 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1887..1905 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1969..1984 /note="100F" primer_bind 2240..2258 /note="372F" primer_bind 2265..2280 /note="100F" protein_bind 2344..2362 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2365..2383 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 2413..2433 /label=TEV /note="TEV-L" misc_feature 2474..2494 /label=TEV /note="TEV-R" CDS 2501..2527 /codon_start=1 /note="FLAG" /translation="DYKDDDDK" polyA_signal 2559..2783 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3067..3114 /bound_moiety="FLP recombinase" /note="FRT" CDS 3122..4144 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4274..4404 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4787..5460) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5605..6465) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6466..6564) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt 1201 gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt 1261 cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct 1321 gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct 1381 gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca 1441 ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt 1501 cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg 1561 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt 1621 ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc 1681 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg 1741 catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct 1801 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtcctag 1861 gacgatctct atcactgata gggagatctc tatcactgat agggagagct ctgcttatat 1921 agacctccca ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag ttgttacgac 1981 attttggaaa gtcccgttga ttttggtgcc aaaacaaact cccattgacg tcaatggggt 2041 ggagacttgg aaatccccgt gagtcaaacc gctatccacg cccattgatg tactgccaaa 2101 accgcatcac catggtaata gcgatgacta atacgtagat gtactgccaa gtaggaaagt 2161 cccataaggt catgtactgg gcataatgcc aggcgggcca tttaccgtca ttgacgtcaa 2221 tagggggcgt acttggcata tgatacactt gatgtactct agaaaaatgt cgtaacaact 2281 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag 2341 ctctccctat cagtgataga gatctcccta tcagtgatag agatcgtcaa gcttggtacc 2401 gagctcggat ccgaaaacct gtacttccaa ggaaccggtg gtcacctgta caatcgatcc 2461 tgcaggagct agcgagaatt tgtattttca gggtgatatc gattacaagg atgacgacga 2521 taagtagctc gaggtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc 2581 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt 2641 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct 2701 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc 2761 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg 2821 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag 2881 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt 2941 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt 3001 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg 3061 tacctagaag ttcctattcc gaagttccta ttctctagaa agtataggaa cttccttggc 3121 caaaaagcct gaactcaccg cgacgtctgt cgagaagttt ctgatcgaaa agttcgacag 3181 cgtctccgac ctgatgcagc tctcggaggg cgaagaatct cgtgctttca gcttcgatgt 3241 aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc gatggtttct acaaagatcg 3301 ttatgtttat cggcactttg catcggccgc gctcccgatt ccggaagtgc ttgacattgg 3361 ggaattcagc gagagcctga cctattgcat ctcccgccgt gcacagggtg tcacgttgca 3421 agacctgcct gaaaccgaac tgcccgctgt tctgcagccg gtcgcggagg ccatggatgc 3481 gatcgctgcg gccgatctta gccagacgag cgggttcggc ccattcggac cgcaaggaat 3541 cggtcaatac actacatggc gtgatttcat atgcgcgatt gctgatcccc atgtgtatca 3601 ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc gcgcaggctc tcgatgagct 3661 gatgctttgg gccgaggact gccccgaagt ccggcacctc gtgcacgcgg atttcggctc 3721 caacaatgtc ctgacggaca atggccgcat aacagcggtc attgactgga gcgaggcgat 3781 gttcggggat tcccaatacg aggtcgccaa catcttcttc tggaggccgt ggttggcttg 3841 tatggagcag cagacgcgct acttcgagcg gaggcatccg gagcttgcag gatcgccgcg 3901 gctccgggcg tatatgctcc gcattggtct tgaccaactc tatcagagct tggttgacgg 3961 caatttcgat gatgcagctt gggcgcaggg tcgatgcgac gcaatcgtcc gatccggagc 4021 cgggactgtc gggcgtacac aaatcgcccg cagaagcgcg gccgtctgga ccgatggctg 4081 tgtagaagta ctcgccgata gtggaaaccg acgccccagc actcgtccga gggcaaagga 4141 atagcacgta ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg 4201 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt 4261 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat 4321 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact 4381 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc 4441 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg 4501 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat 4561 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg 4621 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct 4681 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc 4741 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg 4801 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg 4861 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg 4921 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac 4981 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca 5041 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt 5101 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc 5161 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag 5221 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac 5281 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt 5341 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa 5401 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg 5461 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa 5521 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat 5581 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc 5641 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat 5701 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc 5761 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc 5821 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag 5881 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg 5941 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg 6001 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag 6061 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt 6121 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga 6181 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc 6241 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc 6301 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc 6361 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc 6421 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca 6481 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat 6541 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt 6601 c // >pKK-RNAi-nucEGFPmiR-TEV-mCherry LOCUS Exported 7285 bp ds-DNA circular SYN 26-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucEGFPmiR-TEV-mCHERRY SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7285) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 7285) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..7285 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1138..1852) /codon_start=1 /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELY" misc_feature 1861..2388 /note="BI Promoter" protein_bind 1866..1884 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1887..1905 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1969..1984 /note="100F" primer_bind 2240..2258 /note="372F" primer_bind 2265..2280 /note="100F" protein_bind 2344..2362 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2365..2383 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 2413..2433 /label=TEV /note="TEV-L" misc_feature 2474..2494 /label=TEV /note="TEV-R" CDS 2501..3211 /codon_start=1 /label=mCherry /note="mCherry" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" polyA_signal 3243..3467 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3751..3798 /bound_moiety="FLP recombinase" /note="FRT" CDS 3806..4828 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4958..5088 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(5471..6144) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(6289..7149) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(7150..7248) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt 1201 gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt 1261 cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct 1321 gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct 1381 gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca 1441 ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt 1501 cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg 1561 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt 1621 ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc 1681 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg 1741 catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct 1801 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtcctag 1861 gacgatctct atcactgata gggagatctc tatcactgat agggagagct ctgcttatat 1921 agacctccca ccgtacacgc ctaccgccca tttgcgtcaa tggggcggag ttgttacgac 1981 attttggaaa gtcccgttga ttttggtgcc aaaacaaact cccattgacg tcaatggggt 2041 ggagacttgg aaatccccgt gagtcaaacc gctatccacg cccattgatg tactgccaaa 2101 accgcatcac catggtaata gcgatgacta atacgtagat gtactgccaa gtaggaaagt 2161 cccataaggt catgtactgg gcataatgcc aggcgggcca tttaccgtca ttgacgtcaa 2221 tagggggcgt acttggcata tgatacactt gatgtactct agaaaaatgt cgtaacaact 2281 ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag 2341 ctctccctat cagtgataga gatctcccta tcagtgatag agatcgtcaa gcttggtacc 2401 gagctcggat ccgaaaacct gtacttccaa ggaaccggtg gtcacctgta caatcgatcc 2461 tgcaggagct agcgagaatt tgtattttca gggtgatatc atggtgagca agggcgagga 2521 ggataacatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg agggctccgt 2581 gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg agggcaccca 2641 gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg acatcctgtc 2701 ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca tccccgacta 2761 cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg 2821 cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca tctacaaggt 2881 gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga agaccatggg 2941 ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg gcgagatcaa 3001 gcagaggctg aagctgaagg acggcggcca ctacgacgct gaggtcaaga ccacctacaa 3061 ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagt tggacatcac 3121 ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg gccgccactc 3181 caccggcggc atggacgagc tgtacaagta gctcgaggtt taaacccgct gatcagcctc 3241 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac 3301 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg 3361 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga 3421 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga 3481 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc 3541 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc 3601 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct 3661 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa 3721 acttgattag ggtgatggtt cacgtaccta gaagttccta ttccgaagtt cctattctct 3781 agaaagtata ggaacttcct tggccaaaaa gcctgaactc accgcgacgt ctgtcgagaa 3841 gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga 3901 atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg 3961 cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg ccgcgctccc 4021 gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt gcatctcccg 4081 ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca 4141 gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga cgagcgggtt 4201 cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc 4261 gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc 4321 cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca 4381 cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc 4441 ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt 4501 cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca 4561 tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg gtcttgacca 4621 actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg 4681 cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag 4741 cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc 4801 cagcactcgt ccgagggcaa aggaatagca cgtactacga gatttcgatt ccaccgccgc 4861 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca 4921 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa 4981 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca 5041 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac 5101 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc 5161 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta 5221 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa 5281 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat 5341 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg 5401 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc 5461 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 5521 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 5581 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 5641 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 5701 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 5761 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 5821 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 5881 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 5941 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 6001 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 6061 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 6121 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 6181 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 6241 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 6301 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 6361 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat 6421 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 6481 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg 6541 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 6601 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 6661 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt 6721 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc 6781 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga 6841 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc 6901 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa 6961 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 7021 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 7081 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg 7141 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat 7201 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt 7261 tccccgaaaa gtgccacctg acgtc // >pKK-RNAi-nucCHERRYmiR-TEV-FLAG LOCUS Exported 6598 bp ds-DNA circular SYN 26-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucCHERRYmiR-TEV-FLAG SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6598) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 6598) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..6598 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1144..1852) /codon_start=1 /note="mCherry" /translation="PRTMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRP YEGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERV MNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDG ALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQY ERAEGRHSTGGMDE" misc_feature 1852..2379 /note="BI Promoter" protein_bind 1857..1875 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1878..1896 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1960..1975 /note="100F" primer_bind 2231..2249 /note="372F" primer_bind 2256..2271 /note="100F" protein_bind 2335..2353 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2356..2374 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 2410..2430 /label=TEV /note="TEV-L" misc_feature 2471..2491 /label=TEV /note="TEV-R" CDS 2498..2524 /codon_start=1 /note="FLAG" /translation="DYKDDDDK" polyA_signal 2556..2780 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3064..3111 /bound_moiety="FLP recombinase" /note="FRT" CDS 3119..4141 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4271..4401 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(4784..5457) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(5602..6462) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(6463..6561) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgccg gtggagtggc ggccctcggc gcgttcgtac tgttccacga 1201 tggtgtagtc ctcgttgtgg gaggtgatgt ccaacttgat gttgacgttg taggcgccgg 1261 gcagctgcac gggcttcttg gccttgtagg tggtcttgac ctcagcgtcg tagtggccgc 1321 cgtccttcag cttcagcctc tgcttgatct cgcccttcag ggcgccgtcc tcggggtaca 1381 tccgctcgga ggaggcctcc cagcccatgg tcttcttctg cattacgggg ccgtcggagg 1441 ggaagttggt gccgcgcagc ttcaccttgt agatgaactc gccgtcctgc agggaggagt 1501 cctgggtcac ggtcaccacg ccgccgtcct cgaagttcat cacgcgctcc cacttgaagc 1561 cctcggggaa ggacagcttc aagtagtcgg ggatgtcggc ggggtgcttc acgtaggcct 1621 tggagccgta catgaactga ggggacagga tgtcccaggc gaagggcagg gggccaccct 1681 tggtcacctt cagcttggcg gtctgggtgc cctcgtaggg gcggccctcg ccctcgccct 1741 cgatctcgaa ctcgtggccg ttcacggagc cctccatgtg caccttgaag cgcatgaact 1801 ccttgatgat ggccatgtta tcctcctcgc ccttgctcac catggtccta ggacgatctc 1861 tatcactgat agggagatct ctatcactga tagggagagc tctgcttata tagacctccc 1921 accgtacacg cctaccgccc atttgcgtca atggggcgga gttgttacga cattttggaa 1981 agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg tggagacttg 2041 gaaatccccg tgagtcaaac cgctatccac gcccattgat gtactgccaa aaccgcatca 2101 ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag tcccataagg 2161 tcatgtactg ggcataatgc caggcgggcc atttaccgtc attgacgtca atagggggcg 2221 tacttggcat atgatacact tgatgtactc tagaaaaatg tcgtaacaac tccgccccat 2281 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctcccta 2341 tcagtgatag agatctccct atcagtgata gagatcgtcg gatctaagct tggtaccgag 2401 ctcggatccg aaaacctgta cttccaagga accggtggtc acctgtacaa tcgatcctgc 2461 aggagctagc gagaatttgt attttcaggg tgatatcgat tacaaggatg acgacgataa 2521 gtagctcgag gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc 2581 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct 2641 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg 2701 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg 2761 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta 2821 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt 2881 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct 2941 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg 3001 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtac 3061 ctagaagttc ctattccgaa gttcctattc tctagaaagt ataggaactt ccttggccaa 3121 aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt 3181 ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg 3241 agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta 3301 tgtttatcgg cactttgcat cggccgcgct cccgattccg gaagtgcttg acattgggga 3361 attcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca cgttgcaaga 3421 cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcca tggatgcgat 3481 cgctgcggcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc aaggaatcgg 3541 tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg tgtatcactg 3601 gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat 3661 gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa 3721 caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg aggcgatgtt 3781 cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt tggcttgtat 3841 ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat cgccgcggct 3901 ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg ttgacggcaa 3961 tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat ccggagccgg 4021 gactgtcggg cgtacacaaa tcgcccgcag aagcgcggcc gtctggaccg atggctgtgt 4081 agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg caaaggaata 4141 gcacgtacta cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat 4201 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt 4261 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac 4321 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat 4381 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg 4441 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc 4501 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc 4561 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat 4621 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac 4681 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt 4741 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca 4801 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc 4861 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact 4921 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct 4981 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag 5041 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca 5101 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa 5161 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc 5221 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag 5281 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg 5341 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca 5401 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc 5461 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag 5521 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata 5581 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat 5641 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg 5701 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc 5761 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc 5821 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc 5881 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc 5941 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc 6001 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa 6061 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat 6121 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata 6181 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca 6241 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag 6301 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc 6361 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc 6421 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata 6481 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta 6541 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtc // >pKK-RNAi-nucCHERRYmiR-TEV-EGFP LOCUS Exported 7304 bp ds-DNA circular SYN 28-MAY-2017 DEFINITION Ligation of inverted Fragment 1 into Fragment 2. ACCESSION BI-16_+_RRP6_WT_ VERSION . KEYWORDS pKK-RNAi-nucCHERRYmiR-TEV-EGFP SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7304) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al JOURNAL Unpublished. REFERENCE 2 (bases 1 to 7304) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT SECID/File created by SciEd Central, Scientific & Educational Software SECNOTES|Vector molecule: Fragment 2 Fragment ends: XmaJI and ClaI Fragment size: 7594 Insert molecule: Fragment 1 Fragment ends: XmaJI and ClaI Fragment size: 1494 FEATURES Location/Qualifiers source 1..7304 /organism="synthetic DNA construct" /mol_type="other DNA" misc_feature 270..543 /note="TK poly(A) signal" misc_feature complement(647..705) /label=Neg miR /note="Neg miR" misc_feature complement(785..843) /label=Neg miR /note="Neg miR" misc_feature complement(923..981) /label=Neg miR /note="Neg miR" misc_feature complement(1052..1137) /note="NLS" CDS complement(1144..1852) /codon_start=1 /note="mCherry" /translation="PRTMVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRP YEGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERV MNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDG ALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQY ERAEGRHSTGGMDE" misc_feature 1852..2379 /note="BI Promoter" protein_bind 1857..1875 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 1878..1896 /bound_moiety="Tet repressor" /note="tet operator" primer_bind 1960..1975 /note="100F" primer_bind 2231..2249 /note="372F" primer_bind 2256..2271 /note="100F" protein_bind 2335..2353 /bound_moiety="Tet repressor" /note="tet operator" protein_bind 2356..2374 /bound_moiety="Tet repressor" /note="tet operator" misc_feature 2410..2430 /label=TEV /note="TEV-L" misc_feature 2471..2491 /label=TEV /note="TEV-R" CDS 2498..3217 /codon_start=1 /note="EGFP" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" polyA_signal 3262..3486 /note="BGH poly(A) signal" /note="bovine growth hormone polyadenylation signal" protein_bind 3770..3817 /bound_moiety="FLP recombinase" /note="FRT" CDS 3825..4847 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /note="confers resistance to hygromycin" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 4977..5107 /note="SV40 poly(A) signal" /note="SV40 polyadenylation signal" rep_origin complement(5490..6163) /direction=LEFT /note="pUC origin" /note="pUC origin color: #66cc99; direction: LEFT" CDS complement(6308..7168) /codon_start=1 /product="beta-lactamase" /note="Ampicillin" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(7169..7267) /gene="bla" /note="Amp promoter" ORIGIN 1 gacggatcgg gagatccgct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc 61 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga 121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca 181 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat 241 tctccgctcc atcgttcaga tccttatcga tgctatggca gggcctgccg ccccgacgtt 301 ggctgcgagc cctgggcctt cacccgaact tggggggtgg ggtggggaaa aggaagaaac 361 gcgggcgtat tggccccaat ggggtcacgg tggggtatcg acagagtgcc agccctggga 421 ccgaaccccg cgtttatgaa caaacgaccc aacacccgtg cgttttattc tgtcttttta 481 ttgccgtcat agcgcgggtt ccttccggta ttgtctcctt ccgtgtttca gttagcctcc 541 cccatcagcg aaccgcgggc cctctagatc actagatatg acgcgtgcgg ccgcagatct 601 gggccatttg ttccatgtga gtgctagtaa caggccttgt gtcctgaaat gtactgcgtg 661 gagacgtcag tcagtggcca aaacgtctcc acgcgcagta catttcagca tacagccttc 721 agcaagcctc caggatctgg gccatttgtt ccatgtgagt gctagtaaca ggccttgtgt 781 cctgaaatgt actgcgtgga gacgtcagtc agtggccaaa acgtctccac gcgcagtaca 841 tttcagcata cagccttcag caagcctcca ggatctgggc catttgttcc atgtgagtgc 901 tagtaacagg ccttgtgtcc tgaaatgtac tgcgtggaga cgtcagtcag tggccaaaac 961 gtctccacgc gcagtacatt tcagcataca gccttcagca agcctccagg atcacttaag 1021 actagcgctc caattgcact acctcctctt actacacctt gcgtttcttc tttgggtcga 1081 cctttctctt ctttttaggg tccaccttcc gctttttctt ggggtctgct gcggccttgt 1141 acagctcgtc catgccgccg gtggagtggc ggccctcggc gcgttcgtac tgttccacga 1201 tggtgtagtc ctcgttgtgg gaggtgatgt ccaacttgat gttgacgttg taggcgccgg 1261 gcagctgcac gggcttcttg gccttgtagg tggtcttgac ctcagcgtcg tagtggccgc 1321 cgtccttcag cttcagcctc tgcttgatct cgcccttcag ggcgccgtcc tcggggtaca 1381 tccgctcgga ggaggcctcc cagcccatgg tcttcttctg cattacgggg ccgtcggagg 1441 ggaagttggt gccgcgcagc ttcaccttgt agatgaactc gccgtcctgc agggaggagt 1501 cctgggtcac ggtcaccacg ccgccgtcct cgaagttcat cacgcgctcc cacttgaagc 1561 cctcggggaa ggacagcttc aagtagtcgg ggatgtcggc ggggtgcttc acgtaggcct 1621 tggagccgta catgaactga ggggacagga tgtcccaggc gaagggcagg gggccaccct 1681 tggtcacctt cagcttggcg gtctgggtgc cctcgtaggg gcggccctcg ccctcgccct 1741 cgatctcgaa ctcgtggccg ttcacggagc cctccatgtg caccttgaag cgcatgaact 1801 ccttgatgat ggccatgtta tcctcctcgc ccttgctcac catggtccta ggacgatctc 1861 tatcactgat agggagatct ctatcactga tagggagagc tctgcttata tagacctccc 1921 accgtacacg cctaccgccc atttgcgtca atggggcgga gttgttacga cattttggaa 1981 agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg tggagacttg 2041 gaaatccccg tgagtcaaac cgctatccac gcccattgat gtactgccaa aaccgcatca 2101 ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag tcccataagg 2161 tcatgtactg ggcataatgc caggcgggcc atttaccgtc attgacgtca atagggggcg 2221 tacttggcat atgatacact tgatgtactc tagaaaaatg tcgtaacaac tccgccccat 2281 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctcccta 2341 tcagtgatag agatctccct atcagtgata gagatcgtcg gatctaagct tggtaccgag 2401 ctcggatccg aaaacctgta cttccaagga accggtggtc acctgtacaa tcgatcctgc 2461 aggagctagc gagaatttgt attttcaggg tgatatcatg gtgagcaagg gcgaggagct 2521 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt 2581 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat 2641 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg 2701 cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc 2761 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa 2821 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg 2881 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag 2941 ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat 3001 ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc 3061 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccgccct 3121 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc 3181 cgggatcact ctcggcatgg acgagctgta caagtaatcc ggactcagat ctcgaggttt 3241 aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct 3301 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg 3361 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc 3421 aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat gcggtgggct 3481 ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc cacgcgccct 3541 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg 3601 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg 3661 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac 3721 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtacctag aagttcctat 3781 tccgaagttc ctattctcta gaaagtatag gaacttcctt ggccaaaaag cctgaactca 3841 ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc 3901 agctctcgga gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg 3961 tcctgcgggt aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact 4021 ttgcatcggc cgcgctcccg attccggaag tgcttgacat tggggaattc agcgagagcc 4081 tgacctattg catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg 4141 aactgcccgc tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc 4201 ttagccagac gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat 4261 ggcgtgattt catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg 4321 acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg 4381 actgccccga agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg 4441 acaatggccg cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat 4501 acgaggtcgc caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc 4561 gctacttcga gcggaggcat ccggagcttg caggatcgcc gcggctccgg gcgtatatgc 4621 tccgcattgg tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag 4681 cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta 4741 cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa gtactcgccg 4801 atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa ggaatagcac gtactacgag 4861 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg 4921 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact 4981 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata 5041 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc 5101 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc 5161 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt 5221 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc 5281 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg 5341 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct 5401 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca 5461 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga 5521 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc 5581 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg 5641 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 5701 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt 5761 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 5821 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg 5881 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg 5941 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg 6001 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 6061 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 6121 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 6181 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga 6241 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt 6301 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt 6361 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat 6421 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag 6481 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct 6541 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt 6601 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg 6661 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca 6721 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt 6781 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat 6841 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac 6901 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa 6961 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt 7021 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt 7081 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa 7141 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt 7201 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa 7261 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtc // >pKK-RNAtag-nTER-HA-TEV LOCUS Exported 5315 bp ds-DNA circular SYN 13-OCT-2016 DEFINITION synthetic circular DNA. ACCESSION . VERSION . KEYWORDS pKK-RNAtag-nTER-HA-TEV SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5315) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT LSOWNER| FEATURES Location/Qualifiers source 1..5315 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" CDS 990..1055 /codon_start=1 /note="N terminator RNA BP" /translation="MDAQTRRRERRAEKQAQWKAAN" CDS 1092..1118 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /note="HA" /translation="YPYDVPDYA" misc_feature 1125..1145 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1186..1206 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" polyA_signal 1273..1497 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1781..1828 /bound_moiety="FLP recombinase" /note="FRT" CDS 1836..2858 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2988..3118 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3501..4174) /direction=LEFT /note="pUC ori" CDS complement(4319..5179) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" misc_feature complement(5180..5278) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtacca tggacgcaca aacacgacga cgtgagcgtc 1021 gcgctgagaa acaagctcaa tggaaagctg caaacccacc gctcgaggct gcccaagctt 1081 acaccatggc ctacccatat gatgttccag attacgctgg atccgaaaac ctgtacttcc 1141 aaggaaccgg tggtcacctg tacaatcgat cctgcaggag ctagcgagaa tttgtatttt 1201 cagggtgata tccagcacag tggcggccgc tcgagtctag agggcccgtt taaacccgct 1261 gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc 1321 cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg 1381 catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca 1441 agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt 1501 ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg 1561 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc 1621 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc 1681 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg 1741 accccaaaaa acttgattag ggtgatggtt cacgtaccta gaagttccta ttccgaagtt 1801 cctattctct agaaagtata ggaacttcct tggccaaaaa gcctgaactc accgcgacgt 1861 ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg 1921 agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg 1981 taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg 2041 ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt 2101 gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg 2161 ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga 2221 cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt 2281 tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg 2341 tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg 2401 aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc 2461 gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg 2521 ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg 2581 agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg 2641 gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc 2701 agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg 2761 cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa 2821 accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtactacga gatttcgatt 2881 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga 2941 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg 3001 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt 3061 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta 3121 taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa 3181 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct 3241 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc 3301 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg 3361 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc 3421 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag 3481 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa 3541 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc 3601 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc 3661 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg 3721 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt 3781 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc 3841 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc 3901 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag 3961 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg 4021 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa 4081 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag 4141 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact 4201 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa 4261 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt 4321 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag 4381 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca 4441 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc 4501 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt 4561 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg 4621 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca 4681 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg 4741 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca 4801 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg 4861 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct 4921 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca 4981 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca 5041 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg 5101 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac 5161 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt 5221 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc 5281 cgcgcacatt tccccgaaaa gtgccacctg acgtc // >pKK-RNAtag-3UTR-24MS2SL LOCUS Exported 5306 bp ds-DNA circular SYN 26-MAY-2017 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pKK-RNAtag-3UTR-24MS2SL SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5306) AUTHORS Roman J. Szczesny, Katarzyna Kowalska, Andrzej Dziembowski et al TITLE Direct Submission JOURNAL Exported Sunday, May 28, 2017 from SnapGene 3.3.3 http://www.snapgene.com COMMENT W sekwencji szpilek MS2 jest miesce dla restryktazy NheI. Mozliwe jest klonowanie SLIC z wykorzystaniem miejsca EcoRV. FEATURES Location/Qualifiers source 1..5306 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 232..958 /note="CMV promoter" /note="human cytomegalovirus (CMV) immediate early promoter" protein_bind 820..838 /bound_moiety="repressor" /note="tet operator" protein_bind 841..859 /bound_moiety="repressor" /note="tet operator" misc_feature 1002..1022 /note="TEV-L" /note="enocdes peptide cleaved by TEV protease" misc_feature 1063..1083 /note="TEV-R" /note="enocdes peptide cleaved by TEV protease" stem_loop 1091..1210 /note="MS2 SL" misc_feature 1104..1120 /note="MS2 stem loop" misc_feature 1136..1152 /note="MS2 stem-loop" misc_feature 1167..1183 /note="MS2 stem-loop" polyA_signal 1264..1488 /note="BGH poly(A)signal" /note="bovine growth hormone polyadenylation signal" protein_bind 1772..1819 /bound_moiety="FLP recombinase" /note="FRT" CDS 1827..2849 /codon_start=1 /product="aminoglycoside phosphotransferase from E. coli" /note="Hygromycin B" /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE " polyA_signal 2979..3109 /note="SV40 poly(A)signal" /note="SV40 polyadenylation signal" rep_origin complement(3492..4165) /direction=LEFT /note="pUC ori" CDS complement(4310..5170) /codon_start=1 /product="beta-lactamase" /note="AmpR" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(5171..5275) /gene="bla" /note="AmpR promoter" misc_feature complement(5171..5269) /note="Amp promoter" ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt 241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata 301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc 361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc 421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt 481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt 541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca 601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg 661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc 721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg 781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc 841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga 901 gacgccatcc acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga 961 ctctagcgtt taaacttaag cttggtaccg agctcggatc cgaaaacctg tacttccaag 1021 gaaccggtgg tcacctgtac aatcgatcct gcaggagcta gcgagaattt gtattttcag 1081 ggtgatatcg ggatccgatt accgtacacc atcagggtac gagctagccc atggcgtaca 1141 ccatcagggt acgactagta gatctcgtac accatcaggg tacggaattc tctagagtcg 1201 agttctatag tcgacggatc ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct 1261 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga 1321 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt 1381 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg 1441 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg 1501 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg 1561 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg 1621 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc 1681 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa 1741 aacttgatta gggtgatggt tcacgtacct agaagttcct attccgaagt tcctattctc 1801 tagaaagtat aggaacttcc ttggccaaaa agcctgaact caccgcgacg tctgtcgaga 1861 agtttctgat cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag 1921 aatctcgtgc tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct 1981 gcgccgatgg tttctacaaa gatcgttatg tttatcggca ctttgcatcg gccgcgctcc 2041 cgattccgga agtgcttgac attggggaat tcagcgagag cctgacctat tgcatctccc 2101 gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc 2161 agccggtcgc ggaggccatg gatgcgatcg ctgcggccga tcttagccag acgagcgggt 2221 tcggcccatt cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg 2281 cgattgctga tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt 2341 ccgtcgcgca ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc 2401 acctcgtgca cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag 2461 cggtcattga ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct 2521 tcttctggag gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc 2581 atccggagct tgcaggatcg ccgcggctcc gggcgtatat gctccgcatt ggtcttgacc 2641 aactctatca gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat 2701 gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa 2761 gcgcggccgt ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc 2821 ccagcactcg tccgagggca aaggaatagc acgtactacg agatttcgat tccaccgccg 2881 ccttctatga aaggttgggc ttcggaatcg ttttccggga cgccggctgg atgatcctcc 2941 agcgcgggga tctcatgctg gagttcttcg cccaccccaa cttgtttatt gcagcttata 3001 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc 3061 attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgt ataccgtcga 3121 cctctagcta gagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc 3181 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct 3241 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa 3301 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta 3361 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc 3421 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg 3481 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt 3541 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa 3601 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct 3661 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc 3721 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg 3781 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct 3841 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag 3901 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga 3961 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga 4021 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg 4081 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag 4141 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag 4201 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat 4261 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct 4321 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac 4381 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa 4441 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg 4501 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt 4561 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca 4621 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt 4681 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct 4741 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg 4801 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg 4861 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg 4921 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa 4981 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt 5041 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt 5101 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt 5161 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca 5221 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat 5281 ttccccgaaa agtgccacct gacgtc //