LOCUS       sequence-297922-        5696 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses CtcS protein under the control of T7-lacO promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5696)
  AUTHORS   Jung JK, Alam KK, Verosloff MS, Capdevila DA, Desmau M, Clauer PR, 
            Lee JW, Nguyen PQ, Pasten PA, Matiasek SJ, Gaillard JF, Giedroc DP, 
            Collins JJ, Lucks JB
  TITLE     Cell-free biosensors for rapid detection of water contaminants.
  JOURNAL   Nat Biotechnol. 2020 Jul 6. pii: 10.1038/s41587-020-0571-7. doi: 
            10.1038/s41587-020-0571-7.
  PUBMED    32632301
REFERENCE   2  (bases 1 to 5696)
  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: "Nat 
            Biotechnol. 2020 Jul 6. pii: 10.1038/s41587-020-0571-7. doi: 
            10.1038/s41587-020-0571-7."
COMMENT     Sequence Label: pJBL713
FEATURES             Location/Qualifiers
     source          1..5696
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             15..37
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             560..577
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(586..604)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      600..647
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      684..1139
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(771..790)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     981..1002
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1231..2046)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1231..2046)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     1954..1973
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2038..2057
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2168..2756
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2657..2676
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2910..2927
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2942..3081
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3007..3026)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3167..3189
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3183..3374)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4183..5265)
                     /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     5227..5246
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5266..5343)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5549..5568
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     5653..5672
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5653..5671
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5672..5696
                     /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)."
ORIGIN
        1 cctctagaaa taattttgtt taactttaag aaggagatat accatgaccg acttgagccc
       61 tgccgcagag acattgtcag acattaccac agaacttttc gctgtcaacg gcgcgttatt
      121 acgtgctggt gatgcccttt ccgcccgctt cggtttaact agcgcacgct ggcaggtact
      181 tgggttgtta gcggaggggc ctcaaagtgc ggcccacctg gcacgcgaac gcggtttgcg
      241 tcgtcaagcc gttcagcaaa cggtcgttaa attagtagaa gagggactgg tctcgacttc
      301 acctaatccc gctgatcgtc gcgctccact ggtgagttta accgccaaag gtacagacgc
      361 tttggcacgc attgagcctg ccgagcgctt atggatggag catctggcag ggggtttaga
      421 cccagatgat ttaaaagcta ctctgcgcct gttgcgtgga ttgcgtgctg ttttggccga
      481 gggtttgccc ccggccgccg taggtggcgc agatgctgcc gatgtgaccg gaccggcgaa
      541 gcttgcggcc gcactcgagc accatcacca tcaccattaa ctcgaccgct gagcaataac
      601 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa
      661 ctatatccgg attggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
      721 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
      781 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
      841 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
      901 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
      961 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     1021 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     1081 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     1141 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     1201 tcaaatatgt atccgctcat gaattaattc ttagaaaaac tcatcgagca tcaaatgaaa
     1261 ctgcaattta ttcatatcag gattatcaat accatatttt tgaaaaagcc gtttctgtaa
     1321 tgaaggagaa aactcaccga ggcagttcca taggatggca agatcctggt atcggtctgc
     1381 gattccgact cgtccaacat caatacaacc tattaatttc ccctcgtcaa aaataaggtt
     1441 atcaagtgag aaatcaccat gagtgacgac tgaatccggt gagaatggca aaagtttatg
     1501 catttctttc cagacttgtt caacaggcca gccattacgc tcgtcatcaa aatcactcgc
     1561 atcaaccaaa ccgttattca ttcgtgattg cgcctgagcg agacgaaata cgcgatcgct
     1621 gttaaaagga caattacaaa caggaatcga atgcaaccgg cgcaggaaca ctgccagcgc
     1681 atcaacaata ttttcacctg aatcaggata ttcttctaat acctggaatg ctgttttccc
     1741 ggggatcgca gtggtgagta accatgcatc atcaggagta cggataaaat gcttgatggt
     1801 cggaagaggc ataaattccg tcagccagtt tagtctgacc atctcatctg taacatcatt
     1861 ggcaacgcta cctttgccat gtttcagaaa caactctggc gcatcgggct tcccatacaa
     1921 tcgatagatt gtcgcacctg attgcccgac attatcgcga gcccatttat acccatataa
     1981 atcagcatcc atgttggaat ttaatcgcgg cctagagcaa gacgtttccc gttgaatatg
     2041 gctcataaca ccccttgtat tactgtttat gtaagcagac agttttattg ttcatgacca
     2101 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     2161 gatcttcttg aaatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     2221 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     2281 ctggcttcag cagagcgcag ataccaaata ctgtccttct agtgtagccg tagttaggcc
     2341 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     2401 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     2461 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     2521 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     2581 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     2641 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     2701 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     2761 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     2821 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     2881 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     2941 gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc aatggtgcac
     3001 tctcagtaca atctgctctg atgccgcata gttaagccag tatacactcc gctatcgcta
     3061 cgtgactggg tcatggctgc gccccgacac ccgccaacac ccgctgacgc gccctgacgg
     3121 gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg
     3181 tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca
     3241 gcgtggtcgt gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt
     3301 ttctccagaa gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt
     3361 tcctgtttgg tcactgatgc ctccgtgtaa gggggatttc tgttcatggg ggtaatgata
     3421 ccgatgaaac gagagaggat gctcacgata cgggttactg atgatgaaca tgcccggtta
     3481 ctggaacgtt gtgagggtaa acaactggcg gtatggatgc ggcgggacca gagaaaaatc
     3541 actcagggtc aatgccagcg cttcgttaat acagatgtag gtgttccaca gggtagccag
     3601 cagcatcctg cgatgcagat ccggaacata atggtgcagg gcgctgactt ccgcgtttcc
     3661 agactttacg aaacacggaa accgaagacc attcatgttg ttgctcaggt cgcagacgtt
     3721 ttgcagcagc agtcgcttca cgttcgctcg cgtatcggtg attcattctg ctaaccagta
     3781 aggcaacccc gccagcctag ccgggtcctc aacgacagga gcacgatcat gcgcacccgt
     3841 ggggccgcca tgccggcgat aatggcctgc ttctcgccga aacgtttggt ggcgggacca
     3901 gtgacgaagg cttgagcgag ggcgtgcaag attccgaata ccgcaagcga caggccgatc
     3961 atcgtcgcgc tccagcgaaa gcggtcctcg ccgaaaatga cccagagcgc tgccggcacc
     4021 tgtcctacga gttgcatgat aaagaagaca gtcataagtg cggcgacgat agtcatgccc
     4081 cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg tcgagatccc
     4141 ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc cgctttccag
     4201 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     4261 ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca acagctgatt
     4321 gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg tttgccccag
     4381 caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc tgtcttcggt
     4441 atcgtcgtat cccactaccg agatatccgc accaacgcgc agcccggact cggtaatggc
     4501 gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg gaacgatgcc
     4561 ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt cgccttcccg
     4621 ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag ccagacgcag
     4681 acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt gacccaatgc
     4741 gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa tactgttgat
     4801 gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg cagcttccac
     4861 agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga cgcgttgcgc
     4921 gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta ccatcgacac
     4981 caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa tttgcgacgg
     5041 cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt tgcccgccag
     5101 ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt ccactttttc
     5161 ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg tctgataaga
     5221 gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca ccaccctgaa
     5281 ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc attcgatggt
     5341 gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaagcag cccagtagta
     5401 ggttgaggcc gttgagcacc gccgccgcaa ggaatggtgc atgcaaggag atggcgccca
     5461 acagtccccc ggccacgggg cctgccacca tacccacgcc gaaacaagcg ctcatgagcc
     5521 cgaagtggcg agcccgatct tccccatcgg tgatgtcggc gatataggcg ccagcaaccg
     5581 cacctgtggc gccggtgatg ccggccacga tgcgtccggc gaagcttgcg gccgcactcg
     5641 atcccgcgaa attaatacga ctcactatag gggaattgtg agcggataac aattcc
//
