LOCUS       sequence-297869-        5821 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TetR protein under the control of T7-lacO promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5821)
  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 5821)
  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: pJBL701
FEATURES             Location/Qualifiers
     source          1..5821
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    1..25
                     /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)."
     RBS             40..62
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             69..689
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(142..161)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             711..728
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(737..755)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      751..798
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      835..1290
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(922..941)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1132..1153
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(1382..2197)
                     /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(1382..2197)
                     /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     2105..2124
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     2189..2208
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      2319..2907
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2808..2827
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3061..3078
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    3093..3232
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(3158..3177)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3318..3340
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(3334..3525)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(4334..5416)
                     /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     5378..5397
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(5417..5494)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     5700..5719
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     join(5803..5821,1)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5803..5821
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggaattgtga gcggataaca attcccctct agaaataatt ttgtttaact ttaagaagga
       61 gatataccat gtctagatta gataaaagta aagtgattaa cagcgcatta gagctgctta
      121 atgaggtcgg aatcgaaggt ttaacaaccc gtaaactcgc ccagaagcta ggtgtagagc
      181 agcctacatt gtattggcat gtaaaaaata agcgggcttt gctcgacgcc ttagccattg
      241 agatgttaga taggcaccat actcactttt gccctttaga aggggaaagc tggcaagatt
      301 ttttacgtaa taacgctaaa agttttagat gtgctttact aagtcatcgc gatggagcaa
      361 aagtacattt aggtacacgg cctacagaaa aacagtatga aactctcgaa aatcaattag
      421 cctttttatg ccaacaaggt ttttcactag agaatgcatt atatgcactc agcgctgtgg
      481 ggcattttac tttaggttgc gtattggaag atcaagagca tcaagtcgct aaagaagaaa
      541 gggaaacacc tactactgat agtatgccgc cattattacg acaagctatc gaattatttg
      601 atcaccaagg tgcagagcca gccttcttat tcggccttga attgatcata tgcggattag
      661 agaagcaact taagtgtgaa agtgggtcta agcttgcggc cgcactcgag caccatcacc
      721 atcaccatta actcgaccgc tgagcaataa ctagcataac cccttggggc ctctaaacgg
      781 gtcttgaggg gttttttgct gaaaggagga actatatccg gattggcgaa tgggacgcgc
      841 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
      901 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
      961 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     1021 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     1081 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     1141 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     1201 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     1261 attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa atgtgcgcgg
     1321 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgaattaatt
     1381 cttagaaaaa ctcatcgagc atcaaatgaa actgcaattt attcatatca ggattatcaa
     1441 taccatattt ttgaaaaagc cgtttctgta atgaaggaga aaactcaccg aggcagttcc
     1501 ataggatggc aagatcctgg tatcggtctg cgattccgac tcgtccaaca tcaatacaac
     1561 ctattaattt cccctcgtca aaaataaggt tatcaagtga gaaatcacca tgagtgacga
     1621 ctgaatccgg tgagaatggc aaaagtttat gcatttcttt ccagacttgt tcaacaggcc
     1681 agccattacg ctcgtcatca aaatcactcg catcaaccaa accgttattc attcgtgatt
     1741 gcgcctgagc gagacgaaat acgcgatcgc tgttaaaagg acaattacaa acaggaatcg
     1801 aatgcaaccg gcgcaggaac actgccagcg catcaacaat attttcacct gaatcaggat
     1861 attcttctaa tacctggaat gctgttttcc cggggatcgc agtggtgagt aaccatgcat
     1921 catcaggagt acggataaaa tgcttgatgg tcggaagagg cataaattcc gtcagccagt
     1981 ttagtctgac catctcatct gtaacatcat tggcaacgct acctttgcca tgtttcagaa
     2041 acaactctgg cgcatcgggc ttcccataca atcgatagat tgtcgcacct gattgcccga
     2101 cattatcgcg agcccattta tacccatata aatcagcatc catgttggaa tttaatcgcg
     2161 gcctagagca agacgtttcc cgttgaatat ggctcataac accccttgta ttactgttta
     2221 tgtaagcaga cagttttatt gttcatgacc aaaatccctt aacgtgagtt ttcgttccac
     2281 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     2341 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     2401 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     2461 actgtccttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     2521 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     2581 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     2641 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     2701 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     2761 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     2821 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     2881 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     2941 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
     3001 aaccgtatta ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc
     3061 agcgagtcag tgagcgagga agcggaagag cgcctgatgc ggtattttct ccttacgcat
     3121 ctgtgcggta tttcacaccg caatggtgca ctctcagtac aatctgctct gatgccgcat
     3181 agttaagcca gtatacactc cgctatcgct acgtgactgg gtcatggctg cgccccgaca
     3241 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     3301 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     3361 acgcgcgagg cagctgcggt aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc
     3421 tgcctgttca tccgcgtcca gctcgttgag tttctccaga agcgttaatg tctggcttct
     3481 gataaagcgg gccatgttaa gggcggtttt ttcctgtttg gtcactgatg cctccgtgta
     3541 agggggattt ctgttcatgg gggtaatgat accgatgaaa cgagagagga tgctcacgat
     3601 acgggttact gatgatgaac atgcccggtt actggaacgt tgtgagggta aacaactggc
     3661 ggtatggatg cggcgggacc agagaaaaat cactcagggt caatgccagc gcttcgttaa
     3721 tacagatgta ggtgttccac agggtagcca gcagcatcct gcgatgcaga tccggaacat
     3781 aatggtgcag ggcgctgact tccgcgtttc cagactttac gaaacacgga aaccgaagac
     3841 cattcatgtt gttgctcagg tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc
     3901 gcgtatcggt gattcattct gctaaccagt aaggcaaccc cgccagccta gccgggtcct
     3961 caacgacagg agcacgatca tgcgcacccg tggggccgcc atgccggcga taatggcctg
     4021 cttctcgccg aaacgtttgg tggcgggacc agtgacgaag gcttgagcga gggcgtgcaa
     4081 gattccgaat accgcaagcg acaggccgat catcgtcgcg ctccagcgaa agcggtcctc
     4141 gccgaaaatg acccagagcg ctgccggcac ctgtcctacg agttgcatga taaagaagac
     4201 agtcataagt gcggcgacga tagtcatgcc ccgcgcccac cggaaggagc tgactgggtt
     4261 gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa tgagtgagct aacttacatt
     4321 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4381 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgccagg gtggtttttc
     4441 ttttcaccag tgagacgggc aacagctgat tgcccttcac cgcctggccc tgagagagtt
     4501 gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa atcctgtttg atggtggtta
     4561 acggcgggat ataacatgag ctgtcttcgg tatcgtcgta tcccactacc gagatatccg
     4621 caccaacgcg cagcccggac tcggtaatgg cgcgcattgc gcccagcgcc atctgatcgt
     4681 tggcaaccag catcgcagtg ggaacgatgc cctcattcag catttgcatg gtttgttgaa
     4741 aaccggacat ggcactccag tcgccttccc gttccgctat cggctgaatt tgattgcgag
     4801 tgagatattt atgccagcca gccagacgca gacgcgccga gacagaactt aatgggcccg
     4861 ctaacagcgc gatttgctgg tgacccaatg cgaccagatg ctccacgccc agtcgcgtac
     4921 cgtcttcatg ggagaaaata atactgttga tgggtgtctg gtcagagaca tcaagaaata
     4981 acgccggaac attagtgcag gcagcttcca cagcaatggc atcctggtca tccagcggat
     5041 agttaatgat cagcccactg acgcgttgcg cgagaagatt gtgcaccgcc gctttacagg
     5101 cttcgacgcc gcttcgttct accatcgaca ccaccacgct ggcacccagt tgatcggcgc
     5161 gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag ggccagactg gaggtggcaa
     5221 cgccaatcag caacgactgt ttgcccgcca gttgttgtgc cacgcggttg ggaatgtaat
     5281 tcagctccgc catcgccgct tccacttttt cccgcgtttt cgcagaaacg tggctggcct
     5341 ggttcaccac gcgggaaacg gtctgataag agacaccggc atactctgcg acatcgtata
     5401 acgttactgg tttcacattc accaccctga attgactctc ttccgggcgc tatcatgcca
     5461 taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat ctcgacgctc tcccttatgc
     5521 gactcctgca ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca
     5581 aggaatggtg catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc
     5641 atacccacgc cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg
     5701 gtgatgtcgg cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg
     5761 atgcgtccgg cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag
     5821 g
//
