Table 2.
Primers used in this study.
| Primer | Sequence (5′-3′) | Use |
|---|---|---|
| TgGRA1_cDNA_1F | ACC AGT CGA CTC TAG ATG GTG CGT GTG AGC GCT AT | To clone full length of the gene into XbaI and BamHI sites of the p3XFLAG-CMV-14 plasmid by In-Fusion cloning |
| TgGRA1_cDNA_2R | AGT CAG CCC GGG ATC TCT CTC TCT CTC CTG TTA AGA | |
| TgGRA2_cDNA_1F | ACC AGT CGA CTC TAG ATG TTC GCC GTA AAA CAT TG | |
| TgGRA2_cDNA_2R | AGT CAG CCC GGG ATC TCT GC GAA AAG TCT GGG ACG G | |
| TgGRA3_cDNA_1F | ACCA GTC GAC TCT AGA TGG ACC GTA CCA TAT GTC C | |
| TgGRA3_cDNA_2R | AGT CAG CCC GGG ATC TTT TCT TGG AGG CTT TGT CCA | |
| TgGRA4_cDNA_1F | ACC AGT CGA CTC TAG ATG CAG GGC ACT TGG TTT TC | |
| TgGRA4_cDNA_2R | AGT CAG CCC GGG ATC TCT CTT TGC GCA TTC TTT CCA | |
| TgGRA5_cDNA_1F | ACC AGT CGA CTC TAG ATG GCG TCT GTA AAA CGC GT | |
| TgGRA5_cDNA_2R | AGT CAG CCC GGG ATC TCT CTT CCT CGG CAA CTT CTT | |
| TgGRA6_cDNA_1F | ACC AGT CGA CTC TAG ATG GCA CAC GGT GGC ATC TA | |
| TgGRA6_cDNA_2R | AGT CAG CCC GGG ATC TAA AAT CAA ACT CAT TCA CAC | |
| TgGRA7_cDNA_1F | ACC AGT CGA CTC TAG ATG GCC CGA CAC GCA ATT TT | |
| TgGRA7_cDNA_2R | AGT CAG CCC GGG ATC TCT GGC GGG CAT CCT CCC CAT | |
| TgGRA8_cDNA_1F | ACC AGT CGA CTC TAG ATG GCT TTA CCA TTG CGT GT | |
| TgGRA8_cDNA_2R | AGT CAG CCC GGG ATC TAT TCT GCG TCG TTT GGA CGG | |
| TgGRA9_cDNA_1F | ACC AGT CGA CTCT AGA TGC GGT CAC TCA AGT CAA T | |
| TgGRA9_cDNA_2R | AGT CAG CCC GGG ATC TGA GTC CTC GGT CTT CCT GCG | |
| TgGRA11_212410_cDNA_1F | ACC AGT CGA CTC TAG ATG TCC CGC CGC ATG GCA TC | |
| TgGRA11_212410_cDNA_2R | AGT CAG CCC GGG ATC TTG GCT TCA ACT CGT CCT CTT | |
| TgGRA12_275850_cDNA_1F | ACC AGT CGA CTC TAG ATG GAG ACT GGC CTA AAG GA | |
| TgGRA12_275850_cDNA_2R | AGT CGC CCG GGA TCT CTT CTT TTG TGA AGG TTT C | |
| TgGRA14_cDNA_1F | ACC AGT CGA CTC TAG ATG CAG GCG ATA GCG CGG GG | |
| TgGRA14_cDNA_2R | AGT CAG CCC GGG ATC TTT CGC TTG GTC TCT GGT AGC | |
| TgGRA15_cDNA_1F | ACC AGT CGA CTC TAG ATG GTG ACA ACA ACC ACG CC | |
| TgGRA15_cDNA_2R | AGT CAG CCC GGG ATC TTG GAG TTA CCG CTG ATT GT | |
| TgGRA16_cDNA_1F | ACC AGT CGA CTC TAG ATG TAT CGA AAC CAC TCA GG | |
| TgGRA16_cDNA_2R | AGT CAG CCC GGG ATC TCA TCT GAT CAT TTT TCC GC | |
| TgGRA23_cDNA_1F | ACC AGT CGA CTC TAG ATG GCA GCG CGT GCG GGA AG | |
| TgGRA23_cDNA_2R | AGT CAG CCC GGG ATC TGT TCT TTC GCG CAA GGG GT | |
| TgGRA24_cDNA_1F | ACC AGT CGA CTC TAG ATG CTC CAG ATG GCA CGA TA | |
| TgGRA24_cDNA_2R | AGT CAG CCC GGG ATC TAT TAC CCT TAG TGG GTG GT | |
| TgGRA25_cDNA_1F | ACC AGT CGA CTC TAG ATG AAG CGT TTC TGG TTG TG | |
| TgGRA25_cDNA_2R | AGT CAG CCC GGG ATC TGT TTC TAT CGA ATT CCG GG | |
| TgROP5_cDNA_1F | ACC AGT CGA CTC TAG ATG GCG ACG AAG CTC GCT AG | |
| TgROP5_cDNA_2R | AGT CAG CCC GGG ATC TAG CGA CTG AGG GCG CAG CA | |
| TgROP8_cDNA_1F | ACC AGT CGA CTC TAG ATG TTT TCT GTG TTA CGT AA | |
| TgROP8_cDNA_2R | AGT CAG CCC GGG ATC TTG CCG GTT CTC CAT CAG TT | |
| TgROP9_cDNA_1F | ACC AGT CGA CTC TAG ATG ACG CAC CCA AAT CCC CT | |
| TgROP9_cDNA_2R | AGT CAG CCC GGG ATC TCT GCA TGA TCA ACG AGG GC | |
| TgROP10_cDNA_1F | ACC AGT CGA CTC TAG ATG GGA CGA CCC AGG TGG CC | |
| TgROP10_cDNA_2R | AGT CAG CCC GGG ATC TGT TGG GCG CAT CTT CCG TA | |
| TgROP11_cDNA_1F | ACC AGT CGA CTC TAG ATG TCG TCA TCC AGA TTG GT | |
| TgROP11_cDNA_2R | AGT CAG CCC GGG ATC TCC CCG TGA CGG GGA AGT AC | |
| TgROP12_cDNA_1F | ACC AGT CGA CTC TAG ATG GCA CGC GTT CTT CCT TG | |
| TgROP12_cDNA_2R | AGT CAG CCC GGG ATC TGA ACC GCC TCA AGA GAA AA | |
| TgROP13_cDNA_1F | ACC AGT CGA CTC TAG ATG AAG AGA ACA GAG CTT TG | |
| TgROP13_cDNA_2R | AGT CAG CCC GGG ATC TCA ATA GCC TCA AGG AAT TC | |
| TgROP14_cDNA_1F | ACC AGT CGA CTC TAG ATG TAT TCC TCC CCT CAG TC | |
| TgROP14_cDNA_2R | AGT CAG CCC GGG ATC TCA GCG CTT GCT TCT TCC TA | |
| TgROP15_cDNA_1F | ACC AGT CGA CTC TAG ATG CTG AAA ACG ACA CCT GC | |
| TgROP15_cDNA_2R | AGT CAG CCC GGG ATC TGA AAG GTG AGC TAT GAG GT | |
| TgROP16_cDNA_1F | ACC AGT CGA CTC TAG ATG AAA GTG ACC ACG AAA GG | |
| TgROP16_cDNA_2R | AGT CAG CCC GGG ATC TCA TCC GAT GTG AAG AAA GT | |
| TgROP17_cDNA_1F | ACC AGT CGA CTC TAG ATG GAG TTG GTG TTG TGC TT | |
| TgROP17_cDNA_2R | AGT CAG CCC GGG ATC TCT CCT TCT GTA ATA AAG CC | |
| TgROP18_cDNA_1F | ACC AGT CGA CTC TAG ATG TTT TCG GTA CAG CGG CC | |
| TgROP18_cDNA_2R | AGT CAG CCC GGG ATC TTT CTG TGT GGA GAT GTT CC | |
| TgROP19A_cDNA_1F | ACC AGT CGACTC TAG ATG AGA AGG CGC TGC TTT C | |
| TgROP19A_cDNA_2R | AGT CAG CCC GGG ATC TCT GAG ATC TGG ATG CGC GC | |
| TgROP20_cDNA_1F | ACC AGT CGA CTC TAG ATG CGC CTG GAT GCT GTG TA | |
| TgROP20_cDNA_2R | AGT CAG CCC GGG ATC TGT CAC TTG AAC TTG GCT CC | |
| TgROP23_cDNA_1F | ACC AGT CGA CTC TAG ATG GAA AAG ATC CTG TGG GC | |
| TgROP23_cDNA_2R | AGT CAG CCC GGG ATC TCT TGA TGC CTT TCA ACA GG | |
| TgROP24_cDNA_1F | ACC AGT CGA CTC TAG ATG GCA ACG CGT TCA TTC CT | |
| TgROP24_cDNA_2R | AGT CAG CCC GGG ATC TGG GAT TAC GGG AGA GTG TT | |
| TgROP26_cDNA_1F | ACC AGT CGA CTC TAG ATG TTG TTA AGC ATA TCT GC | |
| TgROP26_cDNA_2R | AGT CAG CCC GGG ATC TTA ATG GGG TAA ACA ACT GC | |
| TgROP34_cDNA_1F | ACC AGT CGA CTC TAG ATG ATG TTT CCT GCC GTC GC | |
| TgROP34_cDNA_2R | AGT CAG CCC GGG ATC TGC TCT CCT GTG CGT CTT CC | |
| TgROP35_cDNA_1F | ACC AGT CGA CTC TAG ATG CCG GAA CAA GAT CTT GC | |
| TgROP35_cDNA_2R | AGT CAG CCC GGG ATC TTT CGT TTT CCT GTT CAT GG | |
| TgROP38_cDNA_1F | ACC AGT CGA CTC TAG ATG AAA AAT ACT CTG TTG TC | |
| TgROP38_cDNA_2R | AGT CAG CCC GGG ATC TAA ATT GAT GCG TTC TTA TC | |
| TgROP39_cDNA_1F | ACC AGT CGA CTC TAG ATG AGC AAA CCT TTT TTC CC | |
| TgROP39_cDNA_2R | AGT CAG CCC GGG ATC TAA CAA TTG ACT CCC GAA GA | |
| TgROP41_cDNA_1F | ACC AGT CGA CTC TAG ATG CGT CAC GTG TTC AAC TC | |
| TgROP41_cDNA_2R | AGT CAG CCC GGG ATC TGG AAA GCA CTT GT GAG GTC | |
| TgGRA7_5UTR_1F | GTG GAT CCC ATG GAG ACA CAC GGT CAA CA | To clone 5′UTR of the TgGRA7 gene into pBS/GFP/HX |
| TgGRA7_5UTR_2R | CGA AGC TTT AAT GCA GCT GTC ATG TCT CG | |
| TgGRA7_3UTR_1F | ATGGGCCCGGTTGGAAAAGGACCCGTATG | To clone 3′UTR of the TgGRA8 gene into pBS/GFP/HX |
| TgGRA7_3UTR_2R | ATGGGCCCACGGAGACTGCCTTGTCTTTC | |
| TgGRA14II_484-gRNA | GAA GTT CTG AGC CGT TTC CTG TTT TAG AGC TAG AAA TAG C | Primer for CRISPR/CAS9 plasmids targeting the TgGRA14 gene (pSAG1::CAS9-U6::sgTgGRA14) |
| TgGRA15(II)_146-gRNA | GCT CGA TAA TTC GGT GGC TTG GGG TTT TAG AGC TAG AAA TAG C | Primer for CRISPR/CAS9 plasmids targeting the TgGRA15 gene (pSAG1::CAS9-U6::sgTgGRA15) |
| Common CAS9-U6-Rv | AAC TTG ACA TCC CCA TTT AC | Common primer for CRISPR/CAS9 plasmids targeting Toxoplasma genes |
| DHFR_GRA14_484_1F | AGG TTC AAG AAG TTC TGA GCC GTT TAA GCT TCG CCA GGC TGT AAA | To amplify an amplicon containing TgGRA14 homology regions surrounding a pyrimethamine-resistant DHFR* cassette |
| DHFR_GRA14_484_2R | CAG ACG CAA CAG AAC CAA GGG GAA TTC ATC CTG CAA GTG CAT AG | |
| DHFR-25ntTgGRA15(II)_146_1F | CAA GTC ACG CTC GAT AAT TCG GTG GAA GCT TCG CCA GGC TGT AAA | To amplify an amplicon containing TgGRA15 homology regions surrounding a pyrimethamine-resistant DHFR* cassette |
| DHFR-21ntTgGRA15(II)_146_2R | GAG CAC CGT AAG ATA CCC AAG GGA ATT CAT CCT GCA AGT GCA TAG | |
| TgGRA7-KOS-1F | CGT CAT GAG TAC CGG GAC AT | To confirm the correct homologous recombination of HXGPRT cassette with the TgGRA7 gene |
| TgGRA7-KOS-2R | ATT CAG ACC TGC TGC GAG CC | |
| TgGRA7-KOS-3R | GCA AGG AAC GAT CAT GCG TG | |
| HXGPRT-KOS-1F | CTTGTCGGGGAGCAACAGCC | |
| TgGRA7_RT_1F | TCA CCA CCA GCA TGG ATA AGG | To confirm the insertion of TgGRA7+UTR cassette into the TgUPRT gene |
| TgGRA7_RT_2R | GCC TCG CTT CCT GAA ATG AAC | |
| TgGRA7+UTR_467-2R | GAT TTT CAG CCA CGC CTG TC | To confirm the insertion of TgGRA7+UTR cassette into the TgUPRT gene |
| TgGRA7+UTR_467-1F | AAG GAC CCG TAT GCA GGT AGC T | |
| TgGRA14screen-1Fv3 | CGA GTT GTA GCT GG CTT TTC | To confirm the insertion of DHFR* cassette into the TgGRA14 gene |
| TgGRA14screen-1Rv3 | TGT CAC GGG GAG ACT AGC GT | |
| TgGRA15(II)_screen_1F | TTT CCA GGA GGA ATC GCG CC | To confirm the insertion of DHFR* cassette into the TgGRA15 gene |
| TgGRA15 (II)_screen_2R | CTG CCT CGT CGT GTT TCC CG | |
| DHFR2-1F | CCA TTG TGA ACA TCC TCA AC | To confirm the insertion of DHFR* cassette into the target gene |
| TgDHFR-TS_screen_2R | CAG ACA CAC CGG TTT CTG CAT | |
| TgGRA7(II)+UTR_1F | ATG CGG CCG CAG GAA AAC AGT GTT TCC GAA | To clone full length of the TgGRA7 containing UTR region into Not1 and EcoR5 sites of the pBluescript+SK plasmid by Iigation cloning (pBluescript-TgGRA7+UTR) |
| TgGRA7(II)+UTR_2R | ACG ATA TCA TGC GTC TTT TGT AGT GAA T | |
| TgGRA14(II)+UTR_1Fv2 | ATT CTA GAA AAT AAT GTG CGC ACA CAA C | To clone full length of the TgGRA14 containing UTR region into Not1 and EcoR5 sites of the pBluescript+SK plasmid by Iigation cloning (pBluescript-TgGRA14+UTR) |
| TgGRA14(II)+UTR_2Rv2 | TCA TCG ATT GCC AGC TCC TTT CAG CTT C | |
| pBlue_UPRT_1F | TGT GGC GTC TCG ATT GTG AGA TAG GGC GAA TTG GAG CTC C | To amplify an amplicon containing TgUPRT homology region surrounding TgGRA14+UTR expressed cassette |
| pBlue_UPRT_2R | TTT CCA TCG ACT CGC CAG CTA GGG AAC AAA AGC TGG GTA C | |
| UpgRNA-1F | GAT CCG CTT CTC TTG TAC TGC | To confirm the insertion of TgGRA14+UTR cassette into the TgUPRT gee |
| DngRNA-2R | AAG CAG GTG CAG CGG ACA AG | |
| CXCL1_RT_1F | CAA TGA GCT GCG CTG TCA GT | Real-time PCR for expression of mouse CXCL1 mRNA |
| CXCL1_RT_2R | TTG AGG TGA ATC CCA GCC AT | |
| CXCL5_RT_1F | CGC TAA TTT GGA GGT GAT CCC | Real-time PCR for expression of mouse CXCL5 mRNA |
| CXCL5_RT_2R | ACT TCC ACC GTA GGG CAC TG | |
| IL1-beta-RT-F1 | CCA AAA GATGAA GGG CTG CT | Real-time PCR for expression of mouse IL-1beta mRNA |
| IL1-beta-RT-R1 | TCA TCT GGA CAG CCC AGG TC | |
| IL6-RT-F1 | TTC CAT CCA GTT GCC TTC TTG | Real-time PCR for expression of mouse IL-6 mRNA |
| IL6-RT-R2 | GAA GGC CGT GGT TGT CAC C | |
| CCL17_RT_1F | ATG TAG GCC GAG AGT GCT GC | Real-time PCR for expression of mouse Ccl17 mRNA |
| CCL17_RT_2R | TGA TAG GA ATG GCC CCT TTG | |
| CCL7_RT_1F | GGA TCT CTG CCA CGC TTC TG | Real-time PCR for expression of mouse CCL7 mRNA |
| CCL7_RT_2R | GGC CCA CAC TTG GAT GCT | |
| LCN2-RT-F1 | CCA GTT CGC CAT GGT ATT TTT C | Real-time PCR for expression of mouse LCN2beta mRA |
| LCN2-RT-R1 | CAC ACT CAC CAC CCA TTC AGT T | |
| CSF3-RT-F1 | CTG GCA GCA GAT GGA AAA CC | Real-time PCR for expression of mouse CSF3 mRNA |
| CSF3-RT-R2 | TGT GTG GGC TGC ACA GTA GG | |
| Ptgs2_RT_1F | ATG TAG GCC GAG AGT GCT GC | Real-time PCR for expression of mouse PTGS2 mRNA |
| Ptgs2_RT_2R | CCA GCA CTT CAC CCA TCA GTT | |
| GAPDH-RT-1F | CCC AGG TCC TCG CTT ATG ATC | Internal control gene for real-time RT-PCR analysis |
| GAPDH-RT-2R | CCT GCT TCA CCA CC TTC TTG AT |