LOCUS       sequence-272862-        8510 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses V. cholerae TniQ, Cas8, Cas7, and Cas6 and crRNA-IS186. 
            The crRNA-IS186 targets IS186 loci in Escherchia coli..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8510)
  AUTHORS   Zhang Y, Sun X, Wang Q, Xu J, Dong F, Yang S, Yang J, Zhang Z, Qian 
            Y, Chen J, Zhang J, Liu Y, Tao R, Jiang Y, Yang J, Yang S
  TITLE     Multicopy chromosomal integration using CRISPR-associated 
            transposases.
  JOURNAL   ACS Synth Biol. 2020 Jun 17. doi: 10.1021/acssynbio.0c00073.
  PUBMED    32551502
REFERENCE   2  (bases 1 to 8510)
  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: "ACS Synth 
            Biol. 2020 Jun 17. doi: 10.1021/acssynbio.0c00073."
COMMENT     Sequence Label: pQCascade-IS186
FEATURES             Location/Qualifiers
     source          1..8510
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(4456..4474)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4470..4517
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(4688..5479)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(5480..5571)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(6632..7714)
                     /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     7676..7695
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(7715..7792)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        7838..7856
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7857..7881
                     /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)."
     promoter        8125..8143
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    8144..8168
                     /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 tactcgccgt caactagcgc ggttgttaga ggtgcagaag tctttcctcg cagtttacta
       61 cggacgcact ccatcccctg ctgtcctttg tgtctgcgag aaaatggcta cgcctcctac
      121 ctttggcact ttcaggggta cgaatactgc cacagccata acgtaccttt aattaccact
      181 tgtagctgtg gtaaggagtt tgactaccga gtatctgggt taaagggcat ttgctgcaaa
      241 tgcaaggagc ctatcacctt aaccagcagg gagaacggtc atgaggcagc gtgtactgtt
      301 tcaaactggc ttgctggcca tgaatctaaa cctctgccaa atcttcctaa aagctaccga
      361 tggggtttag ttcattggtg gatgggtatt aaagatagcg agttcgatca cttttcgttc
      421 gttcaatttt tctcaaactg gccaaggtca ttccactcga taatcgaaga tgaagtagag
      481 ttcaaccttg agcatgctgt tgtcagcacg tctgaattac gactaaaaga tcttcttggt
      541 cgattgtttt tcggttcaat tcggttacct gagcggaatc ttcaacacaa tatcatcctt
      601 ggtgagcttc tctgctattt agaaaatcgt ttatggcaag acaagggatt aatcgccaac
      661 ctcaaaatga acgcgttaga ggcgactgta atgttaaatt gtagcctcga tcagattgca
      721 tcaatggttg aacaacgcat cttgaagcca aatcgaaaaa gcaagcccaa cagccctctt
      781 gatgttaccg attatctatt tcatttcggc gatattttct gtctttggtt agctgagttc
      841 caaagcgatg agtttaaccg ttcgttttat gtgtcgaggt ggtaaatgca aactctgaaa
      901 gaactaatcg catccaatcc tgacgactta acaactgaac ttaagagagc atttcgtcca
      961 ctcacaccgc atattgcaat tgatggtaat gaacttgacg cactgacgat attagtcaat
     1021 ttaaccgata agactgatga tcagaaagac ctgctcgatc gagccaaatg caagcaaaaa
     1081 cttcgagatg aaaaatggtg ggctagctgc ataaattgcg ttaattacag acaaagccat
     1141 aacccaaaat tcccggatat acgttctgaa ggcgtgattc gaacccaagc cctgggtgaa
     1201 ttaccgagtt tcctactctc gtcctcaaaa atcccaccat accattggtc atatagccat
     1261 gattcaaaat acgtcaacaa aagcgcattc ctcaccaatg agttttgttg ggatggtgag
     1321 atctcatgtt tgggtgagct tcttaaagat gcagatcacc cactttggaa tactttgaaa
     1381 aagttaggtt gttctcaaaa aacttgcaaa gcaatggcaa aacaactagc tgatattact
     1441 ctcacgacta tcaatgtcac actcgcacca aattacctga ctcaaatatc tctccccgat
     1501 agtgatacat cctacatttc actctctcct gttgcatcgc tatcgatgca aagccacttt
     1561 catcagaggc tccaagatga aaatcggcat agcgcgataa cgcggtttag ccgaactacc
     1621 aacatggggg tcactgcgat gacatgtggc ggcgcattta gaatgttaaa gtctggcgct
     1681 aagttttcca gcccccctca tcaccgatta aacagtaaac gaagttggtt gacgtcagag
     1741 catgttcagt cattaaaaca gtaccagcgc ctcaacaaaa gcctcatacc tgaaaactct
     1801 cggattgcac tccgtagaaa atacaaaatc gagcttcaaa atatggtcag atcttggttt
     1861 gcaatgcaag accataccct tgattcgaac atacttatcc aacatttgaa tcatgaccta
     1921 tcttacttag gagccacaaa acgttttgca tacgatccag cgatgaccaa gctctttact
     1981 gagcttttga aacgagagtt atcaaattca atcaataatg gtgagcaaca cactaatgga
     2041 tcgtttttag tcctaccgaa tatcagggtt tgtggcgcaa cagctttaag ctccccggta
     2101 acggtgggga ttccatcact tacagctttc tttggcttcg ttcacgcatt tgaacggaat
     2161 ataaatcgca ccacctcatc gtttcgtgtt gaatcctttg cgatatgcgt ccatcaacta
     2221 catgtcgaga agcgaggttt gacagcagag tttgtggaaa aaggcgacgg gactatatcc
     2281 gctcccgcga cccgggatga ctggcagtgt gatgtcgtat ttagccttat tttgaacacc
     2341 aactttgctc aacatattga ccaagatacg ttagttacat cactaccaaa gcgattggct
     2401 cggggttcag caaaaattgc gattgatgac tttaaacata tcaactcatt ctcgacatta
     2461 gaaacagcga tcgaatctct gccaatagaa gctggtaggt ggttatcact ttacgcacag
     2521 tcaaacaata atctaagtga tctattagca gccatgacag aggaccatca gctcatggca
     2581 agctgcgtcg gttaccactt gttagaagag cccaaagata aaccaaactc cctcagaggt
     2641 tacaaacacg ctatcgccga gtgcatcatt ggactcatta actcaatcac ctttagctca
     2701 gagactgatc ccaacacaat cttttggtcg ctaaagaact atcaaaacta cctagtggta
     2761 cagccaagga gtatcaacga tgaaactacc gacaaatcta gcctatgagc gctctatcga
     2821 cccatcagat gtctgttttt ttgtcgtctg gcccgatgat agaaaaacac ctttaaccta
     2881 caattctcgt actctgctcg ggcaaatgga agcggcatca ttagcctatg atgtctcagg
     2941 tcaaccaata aaaagtgcca ccgctgaggc gttagctcaa gggaaccctc atcaagttga
     3001 tttctgccac gttccatacg gtgcgagtca tattgaatgc agtttctccg tctcgttttc
     3061 ttctgaacta cgtcaaccat ataagtgtaa ctcaagcaaa gttaaacaaa cgctagtgca
     3121 attagtcgag ctctacgaaa cgaaaatcgg ctggactgag ctagcaaccc gatatttgat
     3181 gaacatttgc aacggtaaat ggctgtggaa aaatacccgt aaagcctatt gctggaacat
     3241 tgtacttaca ccttggcctt ggaacgggga aaaggttgga tttgaagata ttcgtactaa
     3301 ctacacctca cggcaagact ttaaaaataa taaaaattgg tctgctatag ttgaaatgat
     3361 caaaaccgca ttttctagta ctgatgggct ggcgatattt gaagtcaggg ccaccttgca
     3421 cttgccaacg aatgctatgg tgcggccaag ccaagttttc acagaaaaag aaagtggcag
     3481 taaaagtaaa tctaaaactc aaaacagtcg agtttttcag agtacaacta ttgatggtga
     3541 acgatcgcca atactagggg cctttaaaac gggagcagct attgcaacca ttgacgactg
     3601 gtatcctgaa gccactgagc cactaagggt cggacggttt ggggttcatc gcgaagatgt
     3661 cacttgctac cgtcatccgt ctaccggaaa agattttttc tcgatattac aacaagcaga
     3721 gcactatatt gaagtgttga gcgccaacaa aactcccgct caagaaacta tcaacgacat
     3781 gcacttttta atggctaacc tgattaaggg tgggatgttc cagcataaag gagactgact
     3841 gtgaaatggt attataagac aatcaccttt ctgccagagt tgtgcaacaa cgagtcactg
     3901 gctgcaaagt gtctccgcgt tctgcatgga tttaactatc agtatgagac acgaaatatc
     3961 ggcgtttcat ttccgctttg gtgtgatgca acggttggaa aaaagatttc atttgtcagc
     4021 aagaacaaga tagaactcga cttactactt aaacaacact atttcgtcca aatggaacaa
     4081 cttcaatact ttcatatatc caacactgtt ctcgtcccag aagattgtac atacgtttcc
     4141 tttagacgct gtcaatctat agataagctc acagcagcag ggctggcaag gaaaatcaga
     4201 cgcctggaga aacgtgctct atctagaggc gagcaatttg acccatcatc ttttgctcaa
     4261 aaagagcata ctgcaatagc gcactaccac tcacttgggg agtccagcaa acagacgaac
     4321 cgcaactttc gactcaatat caggatgctc tcggagcaac cccgtgaggg gaactcgatt
     4381 tttagtagct atggcttatc aaattcagaa aactcgtttc agcctgtacc cttaatctga
     4441 cctaggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     4501 tcttgagggg ttttttgctg aaacctcagg catttgagaa gcacacggtc acactgcttc
     4561 cggtagtcaa taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc
     4621 ctgaaccgac gaccgggtca tcgtggccgg atcttgcggc ccctcggctt gaacgaattg
     4681 ttagacatta tttgccgact accttggtga tctcgccttt cacgtagtgg acaaattctt
     4741 ccaactgatc tgcgcgcgag gccaagcgat cttcttcttg tccaagataa gcctgtctag
     4801 cttcaagtat gacgggctga tactgggccg gcaggcgctc cattgcccag tcggcagcga
     4861 catccttcgg cgcgattttg ccggttactg cgctgtacca aatgcgggac aacgtaagca
     4921 ctacatttcg ctcatcgcca gcccagtcgg gcggcgagtt ccatagcgtt aaggtttcat
     4981 ttagcgcctc aaatagatcc tgttcaggaa ccggatcaaa gagttcctcc gccgctggac
     5041 ctaccaaggc aacgctatgt tctcttgctt ttgtcagcaa gatagccaga tcaatgtcga
     5101 tcgtggctgg ctcgaagata cctgcaagaa tgtcattgcg ctgccattct ccaaattgca
     5161 gttcgcgctt agctggataa cgccacggaa tgatgtcgtc gtgcacaaca atggtgactt
     5221 ctacagcgcg gagaatctcg ctctctccag gggaagccga agtttccaaa aggtcgttga
     5281 tcaaagctcg ccgcgttgtt tcatcaagcc ttacggtcac cgtaaccagc aaatcaatat
     5341 cactgtgtgg cttcaggccg ccatccactg cggagccgta caaatgtacg gccagcaacg
     5401 tcggttcgag atggcgctcg atgacgccaa ctacctctga tagttgagtc gatacttcgg
     5461 cgatcaccgc ttccctcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     5521 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa atagctagct
     5581 cactcggtcg ctacgctccg ggcgtgagac tgcggcgggc gctgcggaca catacaaagt
     5641 tacccacaga ttccgtggat aagcagggga ctaacatgtg aggcaaaaca gcagggccgc
     5701 gccggtggcg tttttccata ggctccgccc tcctgccaga gttcacataa acagacgctt
     5761 ttccggtgca tctgtgggag ccgtgaggct caaccatgaa tctgacagta cgggcgaaac
     5821 ccgacaggac ttaaagatcc ccaccgtttc cggcgggtcg ctccctcttg cgctctcctg
     5881 ttccgaccct gccgtttacc ggatacctgt tccgcctttc tcccttacgg gaagtgtggc
     5941 gctttctcat agctcacaca ctggtatctc ggctcggtgt aggtcgttcg ctccaagctg
     6001 ggctgtaagc aagaactccc cgttcagccc gactgctgcg ccttatccgg taactgttca
     6061 cttgagtcca acccggaaaa gcacggtaaa acgccactgg cagcagccat tggtaactgg
     6121 gagttcgcag aggatttgtt tagctaaaca cgcggttgct cttgaagtgt gcgccaaagt
     6181 ccggctacac tggaaggaca gatttggttg ctgtgctctg cgaaagccag ttaccacggt
     6241 taagcagttc cccaactgac ttaaccttcg atcaaaccac ctccccaggt ggttttttcg
     6301 aaggttaagt cagttgggga actgcttaac cttcgatcaa accacctccc caggtggttt
     6361 tttcgtttac agggcaaaag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     6421 tcttttctac tgaaccgctc tagatttcag tgcaatttat ctcttcaaat gtagcacctg
     6481 aagtcagccc catacgatat aagttgtaat tctcatgtta gtcatgcccc gcgcccaccg
     6541 gaaggagctg actgggttga aggctctcaa gggcatcggt cgagatcccg gtgcctaatg
     6601 agtgagctaa cttacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct
     6661 gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     6721 gcgccagggt ggtttttctt ttcaccagtg agacgggcaa cagctgattg cccttcaccg
     6781 cctggccctg agagagttgc agcaagcggt ccacgctggt ttgccccagc aggcgaaaat
     6841 cctgtttgat ggtggttaac ggcgggatat aacatgagct gtcttcggta tcgtcgtatc
     6901 ccactaccga gatgtccgca ccaacgcgca gcccggactc ggtaatggcg cgcattgcgc
     6961 ccagcgccat ctgatcgttg gcaaccagca tcgcagtggg aacgatgccc tcattcagca
     7021 tttgcatggt ttgttgaaaa ccggacatgg cactccagtc gccttcccgt tccgctatcg
     7081 gctgaatttg attgcgagtg agatatttat gccagccagc cagacgcaga cgcgccgaga
     7141 cagaacttaa tgggcccgct aacagcgcga tttgctggtg acccaatgcg accagatgct
     7201 ccacgcccag tcgcgtaccg tcttcatggg agaaaataat actgttgatg ggtgtctggt
     7261 cagagacatc aagaaataac gccggaacat tagtgcaggc agcttccaca gcaatggcat
     7321 cctggtcatc cagcggatag ttaatgatca gcccactgac gcgttgcgcg agaagattgt
     7381 gcaccgccgc tttacaggct tcgacgccgc ttcgttctac catcgacacc accacgctgg
     7441 cacccagttg atcggcgcga gatttaatcg ccgcgacaat ttgcgacggc gcgtgcaggg
     7501 ccagactgga ggtggcaacg ccaatcagca acgactgttt gcccgccagt tgttgtgcca
     7561 cgcggttggg aatgtaattc agctccgcca tcgccgcttc cactttttcc cgcgttttcg
     7621 cagaaacgtg gctggcctgg ttcaccacgc gggaaacggt ctgataagag acaccggcat
     7681 actctgcgac atcgtataac gttactggtt tcacattcac caccctgaat tgactctctt
     7741 ccgggcgcta tcatgccata ccgcgaaagg ttttgcgcca ttcgatggtg tccgggatct
     7801 cgacgctctc ccttatgcga ctcctgcatt aggaaattaa tacgactcac tataggggaa
     7861 ttgtgagcgg ataacaattc ccctgtagaa ataattttgt ttaactttaa taaggagata
     7921 taccatgggt gaactgccga gtaggtagct gataactgga tgctttgcgt gcaaaggaac
     7981 ctgaactcgt gaactgccga gtaggtagct gataacggat ccgaattcga gctcggcgcg
     8041 cctgcaggtc gacaagcttg cggccgcata atgcttaagt cgaacagaaa gtaatcgtat
     8101 tgtacacggc cgcataatcg aaattaatac gactcactat aggggaattg tgagcggata
     8161 acaattcccc atcttagtat attagttaag tataagaagg agatatacat atgtttttgc
     8221 aaagacctaa accttacagc gatgaaagtt tagaaagttt ctttatccga gtggctaaca
     8281 aaaatggcta cggtgatgtc catcgcttcc tagaagccac taaacgattc cttcaagaca
     8341 ttgaccataa tggctatcaa acctttccga ctgatataac tcggataaac ccatactcag
     8401 ctaaaaacag ttccagcgca cgaactgcgt cattcctgaa acttgcacaa ttgacattta
     8461 atgaaccgcc agagctactt gggttggcaa ttaacagaac aaacatgaaa
//
