LOCUS       sequence-192858-        5047 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GST-STh expression plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5047)
  AUTHORS   Zhu Y, Luo Q, Davis SM, Westra C, Vickers TJ, Fleckenstein JM
  TITLE     Molecular determinants of enterotoxigenic Escherichia coli 
            heat-stable toxin secretion and delivery.
  JOURNAL   Infect Immun. 2018 Aug 20. pii: IAI.00526-18. doi: 
            10.1128/IAI.00526-18.
  PUBMED    30126899
REFERENCE   2  (bases 1 to 5047)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Infect 
            Immun. 2018 Aug 20. pii: IAI.00526-18. doi: 10.1128/IAI.00526-18."
COMMENT     Sequence Label: pCW002
FEATURES             Location/Qualifiers
     source          1..5047
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(183..188,189..204,205..211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     primer_bind     869..891
                     /label=pGEX 5'
                     /note="pGEX vectors, Glutathione-S-transferase, forward 
                     primer"
     CDS             918..935
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     primer_bind     complement(1097..1119)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1264..1282)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1350..1454
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1455..1523,1524..2315)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(1673..1692)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2486..3074
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2975..2994
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3228..3245
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        3318..3395
                     /gene="lacI"
                     /label=lacI promoter
     CDS             3396..4478
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(3415..3434)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(4527..4532,4533..4550,4551..4557)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4565..4581
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     4589..4605
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4589..4605
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(4601..5047,1..81)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(4621..4638)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4621..4637)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4630..4652)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
ORIGIN
        1 agcttatcga ctgcacggtg caccaatgct tctggcgtca ggcagccatc ggaagctgtg
       61 gtatggctgt gcaggtcgta aatcactgca taattcgtgt cgctcaaggc gcactcccgt
      121 tctggataat gttttttgcg ccgacatcat aacggttctg gcaaatattc tgaaatgagc
      181 tgttgacaat taatcatcgg ctcgtataat gtgtggaatt gtgagcggat aacaatttca
      241 cacaggaaac agtattcatg tcccctatac taggttattg gaaaattaag ggccttgtgc
      301 aacccactcg acttcttttg gaatatcttg aagaaaaata tgaagagcat ttgtatgagc
      361 gcgatgaagg tgataaatgg cgaaacaaaa agtttgaatt gggtttggag tttcccaatc
      421 ttccttatta tattgatggt gatgttaaat taacacagtc tatggccatc atacgttata
      481 tagctgacaa gcacaacatg ttgggtggtt gtccaaaaga gcgtgcagag atttcaatgc
      541 ttgaaggagc ggttttggat attagatacg gtgtttcgag aattgcatat agtaaagact
      601 ttgaaactct caaagttgat tttcttagca agctacctga aatgctgaaa atgttcgaag
      661 atcgtttatg tcataaaaca tatttaaatg gtgatcatgt aacccatcct gacttcatgt
      721 tgtatgacgc tcttgatgtt gttttataca tggacccaat gtgcctggat gcgttcccaa
      781 aattagtttg ttttaaaaaa cgtattgaag ctatcccaca aattgataag tacttgaaat
      841 ccagcaagta tatagcatgg cctttgcagg gctggcaagc cacgtttggt ggtggcgacc
      901 atcctccaaa atcggatctg gttccgcgtg gatccgatcc ccgggtaccg agctcgaata
      961 gtagcaatta ctgctgtgaa ttgtgttgta atcctgcttg taccgggtgc tattgagaat
     1021 tcccgggtcg actcgagcgg ccgcatcgtg actgactgac gatctgcctc gcgcgtttcg
     1081 gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca gcttgtctgt
     1141 aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc
     1201 ggggcgcagc catgacccag tcacgtagcg atagcggagt gtataattct tgaagacgaa
     1261 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     1321 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     1381 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     1441 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     1501 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     1561 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     1621 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     1681 gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt
     1741 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     1801 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     1861 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     1921 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     1981 gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta actggcgaac
     2041 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     2101 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     2161 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     2221 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     2281 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     2341 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     2401 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     2461 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     2521 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     2581 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     2641 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     2701 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     2761 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     2821 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     2881 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     2941 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     3001 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     3061 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     3121 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     3181 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     3241 gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     3301 cacaccgcat aaattccgac accatcgaat ggcgcaaaac ctttcgcggt atggcatgat
     3361 agcgcccgga agagagtcaa ttcagggtgg tgaatgtgaa accagtaacg ttatacgatg
     3421 tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac caggccagcc
     3481 acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg aattacattc
     3541 ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc gttgccacct
     3601 ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct cgcgccgatc
     3661 aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa gcctgtaaag
     3721 cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac tatccgctgg
     3781 atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg ttatttcttg
     3841 atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac ggtacgcgac
     3901 tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta gcgggcccat
     3961 taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc actcgcaatc
     4021 aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt tttcaacaaa
     4081 ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc aacgatcaga
     4141 tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt gcggatatct
     4201 cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg ttaaccacca
     4261 tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg caactctctc
     4321 agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa agaaaaacca
     4381 ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     4441 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtaagt
     4501 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
     4561 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacggattc
     4621 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     4681 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     4741 cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc tttgcctggt ttccggcacc
     4801 agaagcggtg ccggaaagct ggctggagtg cgatcttcct gaggccgata ctgtcgtcgt
     4861 cccctcaaac tggcagatgc acggttacga tgcgcccatc tacaccaacg tgacctatcc
     4921 cattacggtc aatccgccgt ttgttcccac ggagaatccg acgggttgtt actcgctcac
     4981 atttaatgtt gatgaaagct ggctacagga aggccagacg cgaattattt ttgatggcgt
     5041 tggaatt
//
