TABLE 4.
Primers used for construction of unmarked deletion mutants and plasmids for heterologous expression
| Name | Sequencea | Restriction site |
|---|---|---|
| upfor_Chol11_nov2c218 | TTTTTTTTCTAGACGTATCGCCGTGATGAGGAT | XbaI |
| uprev_Chol11_nov2c218 | CCGCGATGGTTTGAGGAGAG | |
| dnfor_Chol11_nov2c218 | CTCTCCTCAAACCATCGCGG GGCTCTCTTCCTTTTCGTTC | |
| dnrev_Chol11_nov2c218 | TTTTTTTAAGCTTGTATCTTCCGGGCCGATCAT | HindIII |
| upfor_Chol11_nov2c219-220 | TTTTTTTCTCGAGATCCTGCAGTCAATCGGTCG | XhoI |
| uprev_Chol11_nov2c219-220 | GCGGGAACGCCTGAACGGAACGAAAAGGAAGAGAGCC | |
| dnfor_Chol11_nov2c219-220 | CGTTCAGGCGTTCCCGC | |
| dnrev_Chol11_nov2c219-220 | TTTTTTTTCTAGATGCCCATATCTCCGACTGGA | XbaI |
| upfor_Chol11_scd4A | TTTTTTTCTCGAGCATCATCAGGACGTCGGTGT | XhoI |
| uprev_Chol11_ scd4A | ACGGTGGACGAGGAAGATGC | |
| dnfor_Chol11_ scd4A | GCATCTTCCTCGTCCACCGTGCCGCTTCTCCCAGGATCAT | |
| dnrev_Chol11_ scd4A | TTTTTTTCTAGATTAGTGTGACGAGCCAGACG | XbaI |
| upfor_Chol1_kshA | TTTTTTTTCTAGATTCGGTGGATTGGGTCGAAG | XbaI |
| uprev_Chol1_kshA | GTCGTTACCCTCAGGCGTTC | |
| dnfor_Chol1_kshA | GAACGCCTGAGGGTAACGACAGGATAACGCTGTGCCAGAC | |
| dnrev_Chol1_kshA | TTTTTTTAAGCTTTCAATCCAAGCCTACGGACG | HindIII |
| expfor_Chol11_nov2c218 | TTTTTTTCTCGAGATGGCGCTGTTGGATGGAC | XhoI |
| exprev_Chol11_nov2c218 | TTTTTTTTCTAGATCACGCAAAAGCCGTCCC | XbaI |
| expfor_Chol11_nov2c219-220 | TTTTTTTCTCGAGGTGGACGAGGAAGATGCCTTTG | XhoI |
| exprev_Chol11_nov2c219-220 | TTTTTTTTCTAGATCAGTCCAGCAGCAGATGCA | XbaI |
| expfor_Chol11_nov2c221 | TTTTTTTCTCGAGATGAAACTCGGATTTTCCCCGG | XhoI |
| exprev_Chol11_nov2c221 | TTTTTTTTCTAGATCAGGCGTTCCCGCG | XbaI |
| expfor_Chol11_nov2c221-222 | TTTTTTTCTCGAGATGGAATTCGCATTTACCGACGAACAACAG | XhoI |
| exprev_Chol11_nov2c221-222 | TTTTTTTTCTAGATCAGGCGTTCCCGCGCG | XbaI |
| expfor_Chol1_scd1A | TTTTTTTCTCGAGATGAGCACTATCGAGCAGTT | XhoI |
| exprev_Chol1_scd1A | TTTTTTTTCTAGATTACTTGCTATCCGGCAGGC | XbaI |
| expfor_Chol1_scd1AB | TTTTTTTAAGCTTATGAGCACTATCGAGCAGTT | HindIII |
| exprev_Chol1_scd1AB | TTTTTTTTCTAGATCAGTCCAGCGTCTTGC | XbaI |
| expfor _Chol1_kshAChol1 | TTTTTTTCTCGAGGAACGCCTGAGGGTAACGAC | XhoI |
| exprev_Chol1_kshAChol1 | TTTTTTTGGATCCTCATTTGCGCTGGTCGACC | BamHI |
| expfor_Chol11_nov2c066 | TTTTTTTCTCGAGCGATGGAGAGGCAGGACTCG | XhoI |
| exprev_Chol11_nov2c066 | TTTTTTTGGATCCTCAATCGACCCCCGGCG | BamHI |
| expfor_Chol11_nov2c228 | TTTTTTTCTCGAGAATCTGACGGAGCCTTTTCC | XhoI |
| exprev_Chol11_nov2v228 | TTTTTTTGGATCCTTATTCCGCGGCCGC | BamHI |
| expfor_Chol11_nov2c407 | TTTTTTTCTCGAGCGCAGTGAGAGGATCAAGCA | XhoI |
| exprev_Chol11_nov2c407 | TTTTTTTGGATCCTTATGCCGCCTGCGTGG | BamHI |
| expfor_Chol11_nov2c430 | TTTTTTTCTCGAGACAAGAGAGAGGATGGACGG | XhoI |
| exprev_Chol11_nov2c430 | TTTTTTTGGATCCTCATTCGGCCGCTGC | BamHI |
| expfor_Chol11_nov2c440 | TTTTTTTCTCGAGGCGAGTGAGAGGATCAAGCA | XhoI |
| exprev_Chol11_nov2c440 | TTTTTTTGGATCCTTATGCCGCCTGCGTGG | BamHI |
| expfor_Chol1_kshBChol1 | TTTTTTTGGATCCTTCGTCATGAGTATCTGAGC | BamHI |
| exprev_Chol1_kshBChol1 | TTTTTTTTCTAGATCAGTCGGGAAAGCG | XbaI |
| expfor_Chol11_novbp123 | TTTTTTTGGATCCCGCACACTAAGGGGATGAGG | BamHI |
| exprev_Chol11_novbp123 | TTTTTTTTCTAGATCAGGCTTCGACCACGAG | XbaI |
| M13 for (–43) | AGGGTTTTCCCAGTCACGACGTT | |
| M13 rev (–49) | GAGCGGATAACAATTTCACACAGG | |
| pEX18AP_backbone_for | AGGAGACATGAACGATGAACA | |
| pEX18AP_backbone_rev | TTTTTTTCCCGGGTCGGCATTTTCTTTTGCGTT | |
| pDM4_MCS_for | ACTTAACGGCTGACATGGGA | |
| pDM4_MCS_rev | GCGAAGTGATCTTCCGTCAC | |
| pDM4_backbone_for | AAGATGTGGCGTGTTACGGT | |
| pDM4_backbone_rev | AGGCTCTGGGAGGCAGAATA |
a
Underlined, restriction sites.