LOCUS       sequence-305681-       10213 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Clostridial vector, encoding a targetron carrying a lox71 site in 
            the reverse (antisense) orientation. Retarget intron to insert lox71
            onto genome at desired site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10213)
  AUTHORS   Cerisy T, Rostain W, Chhun A, Boutard M, Salanoubat M, Tolonen AC
  TITLE     A Targetron-Recombinase System for Large-Scale Genome Engineering of
            Clostridia.
  JOURNAL   mSphere. 2019 Dec 11;4(6). pii: 4/6/e00710-19. doi: 
            10.1128/mSphere.00710-19.
  PUBMED    31826971
REFERENCE   2  (bases 1 to 10213)
  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: "mSphere."; 
            date: "2019-12-11"; volume: "4(6). pii: 4/6/e00710-19. doi"; pages: 
            " 10.1128/mSphere.00710-19"
COMMENT     Sequence Label: pQlox71R
FEATURES             Location/Qualifiers
     source          1..10213
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1944..1960)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2121..2137)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(2463..2834)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            2867..2976
                     /note="incP origin of transfer"
     rep_origin      7625..8213
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8114..8133
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8367..8384
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8501..8522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8537..8542,8543..8560,8561..8567)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8575..8591
                     /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     8580..8602
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8599..8615
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8599..8615
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     protein_bind    9805..9838
                     /label=lox71
                     /bound_moiety="Cre recombinase"
                     /note="Left element (LE) mutant of loxP (Araki et al., 
                     2010). Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     9940..9959
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        9940..9958
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             10121..10143
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
ORIGIN
        1 tgcgttgatg caatttctat gcactcgtag tagtctgaga agggtaacgc cctttacatg
       61 gcaaaggggt acagttattg tgtactaaaa ttaaaaattg attagggagg aaaacctcaa
      121 aatgaaacca acaatggcaa ttttagaaag aatcagtaaa aattcacaag aaaatataga
      181 cgaagttttt acaagacttt atcgttatct tttacgtcca gatatttatt acgtggcgta
      241 tcaaaattta tattccaata aaggagcttc cacaaaagga atattagatg atacagcgga
      301 tggctttagt gaagaaaaaa taaaaaagat tattcaatct ttaaaagacg gaacttacta
      361 tcctcaacct gtacgaagaa tgtatattgc aaaaaagaat tctaaaaaga tgagaccttt
      421 aggaattcca actttcacag ataaattgat ccaagaagct gtgagaataa ttcttgaatc
      481 tatctatgaa ccggtattcg aagatgtgtc tcacggtttt agacctcaac gaagctgtca
      541 cacagctttg aaaacaatca aaagagagtt tggcggcgca agatggtttg tggagggaga
      601 tataaaaggc tgcttcgata atatagacca cgttacactc attggactca tcaatcttaa
      661 aatcaaagat atgaaaatga gccaattgat ttataaattt ctaaaagcag gttatctgga
      721 aaactggcag tatcacaaaa cttacagcgg aacacctcaa ggtggaattc tatctcctct
      781 tttggccaac atctatcttc atgaattgga taagtttgtt ttacaactca aaatgaagtt
      841 tgaccgagaa agtccagaaa gaataacacc tgaatatcgg gagctccaca atgagataaa
      901 aagaatttct caccgtctca agaagttgga gggtgaagaa aaagctaaag ttcttttaga
      961 atatcaagaa aaacgtaaaa gattacccac actcccctgt acctcacaga caaataaagt
     1021 attgaaatac gtccggtatg cggacgactt cattatctct gttaaaggaa gcaaagagga
     1081 ctgtcaatgg ataaaagaac aattaaaact ttttattcat aacaagctaa aaatggaatt
     1141 gagtgaagaa aaaacactca tcacacatag cagtcaaccc gctcgttttc tgggatatga
     1201 tatacgagta aggagatctg gaacgataaa acgatctggt aaagtcaaaa agagaacact
     1261 caatgggagt gtagaactcc ttattcctct tcaagacaaa attcgtcaat ttatttttga
     1321 caagaaaata gctatccaaa agaaagatag ctcatggttt ccagttcaca ggaaatatct
     1381 tattcgttca acagacttag aaatcatcac aatttataat tctgaactcc gcgggatttg
     1441 taattactac ggtctagcaa gtaattttaa ccagctcaat tattttgctt atcttatgga
     1501 atacagctgt ctaaaaacga tagcctccaa acataaggga acactttcaa aaaccatttc
     1561 catgtttaaa gatggaagtg gttcgtgggg gatcccgtat gagataaagc aaggtaagca
     1621 gcgccgttat tttgcaaatt ttagtgaatg taaatcccct tatcaattta cggatgagat
     1681 aagtcaagct cctgtattgt atggctatgc ccggaatact cttgaaaaca ggttaaaagc
     1741 taaatgttgt gaattatgtg ggacgtctga tgaaaatact tcctatgaaa ttcaccatgt
     1801 caataaggtc aaaaatctta aaggcaaaga aaaatgggaa atggcaatga tagcgaaaca
     1861 acgtaaaact cttgttgtat gctttcattg tcatcgtcac gtgattcata aacacaagtg
     1921 agatgggtac cctcgaggaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac
     1981 cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat
     2041 agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg
     2101 cgggtaccga gctcgaattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct
     2161 ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc
     2221 gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc
     2281 ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat aggggatcgg
     2341 tcttgccttg ctcgtcggtg atgtacttca ccagctccgc gaagtcgctc ttcttgatgg
     2401 agcgcatggg gacgtgcttg gcaatcacgc gcaccccccg gccgttttag cggctaaaaa
     2461 agtcatggct ctgccctcgg gcggaccacg cccatcatga ccttgccaag ctcgtcctgc
     2521 ttctcttcga tcttcgccag cagggcgagg atcgtggcat caccgaaccg cgccgtgcgc
     2581 gggtcgtcgg tgagccagag tttcagcagg ccgcccaggc ggcccaggtc gccattgatg
     2641 cgggccagct cgcggacgtg ctcatagtcc acgacgcccg tgattttgta gccctggccg
     2701 acggccagca ggtaggccga caggctcatg ccggccgccg ccgccttttc ctcaatcgct
     2761 cttcgttcgt ctggaaggca gtacaccttg ataggtgggc tgcccttcct ggttggcttg
     2821 gtttcatcag ccatccgctt gccctcatct gttacgccgg cggtagccgg ccagcctcgc
     2881 agagcaggat tcccgttgag caccgccagg tgcgaataag ggacagtgaa gaaggaacac
     2941 ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct gacgccgttg gatacaccaa
     3001 ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc gtgcgaaaaa ggatggatat
     3061 accgaaaaaa tcgctataat gaccccgaag cagggttatg cagcggaaaa gatcccccga
     3121 agcaaactta agagtgtgtt gatagtgcag tatcttaaaa ttttgtgtat aataggaatt
     3181 gaagttaaat tagatgctaa aaatttgtaa ttaagaagga gggattcgtc atgttggtat
     3241 tccaaatgcg taatgtagat aaaacatcta ctgttttgaa acagactaaa aacagtgatt
     3301 acgcagataa ataaatacgt tagattaatt cctaccagtg actaatctta tgacttttta
     3361 aacagataac taaaattaca aacaaatcgt ttaacttctg tatttattta cagatgtaat
     3421 cacttcagga gtaattacat gaacaaaaat ataaaatatt ctcaaaactt tttaacgagt
     3481 gaaaaagtac tcaaccaaat aataaaacaa ttgaatttaa aagaaaccga taccgtttac
     3541 gaaattggaa caggtaaagg gcatttaacg acgaaactgg ctaaaataag taaacaggta
     3601 acgtctattg aattagacag tcatctattc aacttatcgt cagaaaaatt aaaactgaac
     3661 attcgtgtca ctttaattca ccaagatatt ctacagtttc aattccctaa caaacagagg
     3721 tataaaattg ttgggagtat tccttaccat ttaagcacac aaattattaa aaaagtggtt
     3781 tttgaaagcc atgcgtctga catctatctg attgttgaag aaggattcta caagcgtacc
     3841 ttggatattc accgaacact agggttgctc ttgcacactc aagtctcgat tcagcaattg
     3901 cttaagctgc cagcggaatg ctttcatcct aaaccaaaag taaacagtgt cttaataaaa
     3961 cttacccgcc ataccacaga tgttccagat aaatattgga agctatatac gtactttgtt
     4021 tcaaaatggg tcaatcgaga atatcgtcaa ctgtttacta aaaatcagtt tcatcaagca
     4081 atgaaacacg ccaaagtaaa caatttaagt accattactt atgagcaagt attgtctatt
     4141 tttaatagtt atctattatt taacgggagg aaataattct atgagtcgct tttttaaatt
     4201 tggaaagtta cacgttacta aagggaatgg agataaatta ttagatatac tactgacagc
     4261 ttccaagaag ctaaagaggt ccctagcgcc tacggggaat ttgggcaaat caaaagaaaa
     4321 attctcccaa gaattagaaa tgagtagatc aaattattca cgaatagaat caggaaaatc
     4381 agatccaacc ataaaaacac tagaacaaat tgcaaagtta actaactcaa cgctagtagt
     4441 ggatttaatc ccaaatgagc caacagaacc agaaccagaa acagaatcag aacaagtaac
     4501 attggattta gaaatggaag aagaaaaaag caatgacttc gtgtgaataa tgcacgaaat
     4561 cgttgcttat tttttttaaa agcggtatac tagatataac gaaacaacga actgaataga
     4621 aacgaaaaaa gagccatgac acatttataa aatgtttgac gacattttat aaatgcatag
     4681 cccgataaga ttgccaaacc aacgcttatc agttagtcag atgaactctt ccctcgtaag
     4741 aagttattta attaactttg tttgaagacg gtatataacc gtactatcat tatataggga
     4801 aatcagagag ttttcaagta tctaagctac tgaatttaag aattgttaag caatcaatcg
     4861 gaaatcgttt gattgctttt tttgtattca tttatagaag gtggagtttg tatgaatcat
     4921 gatgaatgta aaacttatat aaaaaatagt ttattggaga taagaaaatt agcaaatatc
     4981 tatacactag aaacgtttaa gaaagagtta gaaaagagaa atatctactt agaaacaaaa
     5041 tcagataagt atttttcttc ggagggggaa gattatatat ataagttaat agaaaataac
     5101 aaaataattt attcgattag tggaaaaaaa ttgacttata aaggaaaaaa atctttttca
     5161 aaacatgcaa tattgaaaca gttgaatgaa aaagcaaacc aagttaatta aacaacctat
     5221 tttataggat ttataggaaa ggagaacagc tgaatgaata tcccttttgt tgtagaaact
     5281 gtgcttcatg acggcttgtt aaagtacaaa tttaaaaata gtaaaattcg ctcaatcact
     5341 accaagccag gtaaaagcaa aggggctatt tttgcgtatc gctcaaaatc aagcatgatt
     5401 ggcggtcgtg gtgttgttct gacttccgag gaagcgattc aagaaaatca agatacattt
     5461 acacattgga cacccaacgt ttatcgttat ggaacgtatg cagacgaaaa ccgttcatac
     5521 acgaaaggac attctgaaaa caatttaaga caaatcaata ccttctttat tgattttgat
     5581 attcacacgg caaaagaaac tatttcagca agcgatattt taacaaccgc tattgattta
     5641 ggttttatgc ctactatgat tatcaaatct gataaaggtt atcaagcata ttttgtttta
     5701 gaaacgccag tctatgtgac ttcaaaatca gaatttaaat ctgtcaaagc agccaaaata
     5761 atttcgcaaa atatccgaga atattttgga aagtctttgc cagttgatct aacgtgtaat
     5821 cattttggta ttgctcgcat accaagaacg gacaatgtag aattttttga tcctaattac
     5881 cgttattctt tcaaagaatg gcaagattgg tctttcaaac aaacagataa taagggcttt
     5941 actcgttcaa gtctaacggt tttaagcggt acagaaggca aaaaacaagt agatgaaccc
     6001 tggtttaatc tcttattgca cgaaacgaaa ttttcaggag aaaagggttt aatagggcgt
     6061 aataacgtca tgtttaccct ctctttagcc tactttagtt caggctattc aatcgaaacg
     6121 tgcgaatata atatgtttga gtttaataat cgattagatc aacccttaga agaaaaagaa
     6181 gtaatcaaaa ttgttagaag tgcctattca gaaaactatc aaggggctaa tagggaatac
     6241 attaccattc tttgcaaagc ttgggtatca agtgatttaa ccagtaaaga tttatttgtc
     6301 cgtcaagggt ggtttaaatt caagaaaaaa agaagcgaac gtcaacgtgt tcatttgtca
     6361 gaatggaaag aagatttaat ggcttatatt agcgaaaaaa gcgatgtata caagccttat
     6421 ttagtgacga ccaaaaaaga gattagagaa gtgctaggca ttcctgaacg gacattagat
     6481 aaattgctga aggtactgaa ggcgaatcag gaaattttct ttaagattaa accaggaaga
     6541 aatggtggca ttcaacttgc tagtgttaaa tcattgttgc tatcgatcat taaagtaaaa
     6601 aaagaagaaa aagaaagcta tataaaggcg ctgacaaatt cttttgactt agagcataca
     6661 ttcattcaag agactttaaa caagctagca gaacgcccta aaacggacac acaactcgat
     6721 ttgtttagct atgatacagg ctgaaaataa aacccgcact atgccattac atttatatct
     6781 atgatacgtg tttgtttttt ctttgctgtt tagcgaatga ttagcagaaa tatacagagt
     6841 aagattttaa ttaattatta gggggagaag gagagagtag cccgaaaact tttagttggc
     6901 ttggactgaa cgaagtgagg gaaaggctac taaaacgtcg aggggcagtg agagcgaagc
     6961 gaacacttga ttttttaatt ttctatcttt tataggtcat tagagtatac ttatttgtcc
     7021 tataaactat ttagcagcat aatagattta ttgaataggt catttaagtt gagcatatta
     7081 gaggaggaaa atcttggaga aatatttgaa gaacccgatt acatggattg gattagttct
     7141 tgtggttacg tggtttttaa ctaaaagtag tgaatttttg atttttggtg tgtgtgtctt
     7201 gttgttagta tttgctagtc aaagtgatta aatagaattg accacttctg cgctcggccc
     7261 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     7321 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     7381 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     7441 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     7501 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     7561 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     7621 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     7681 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     7741 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     7801 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     7861 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     7921 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     7981 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     8041 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     8101 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     8161 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     8221 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     8281 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     8341 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     8401 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     8461 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     8521 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     8581 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag cttgcatgcc
     8641 tgcagtttgg tcaacattta acctctaatc ttctagttca tttatatgtc catgaattga
     8701 atttcataca tagttttcaa ttgtatggat gtagtatgac agaaaatcaa aaaatcatac
     8761 ttattatatt taggtagtta ctgttaattt ttagtatgtt aatttctaat ctgagttctt
     8821 tccgtattat aataagtgta aagatcacac catattttta cagtttttac acgtttcatt
     8881 aataatttct tatatttctt tatttgtttg taaaatttac ttaaatttcg ccagaaaaca
     8941 aaagaaagcc tttactaatt aatagtttag tgatactctt ttatgtaggt attttttaaa
     9001 atacattaaa cctaggtaat tgaggaaagt tacaattacc attatataag gaggatattc
     9061 aaaccatatg ggatccccag cttcgtcgat cgtgaagatt taatcaatgt gcgcccagat
     9121 agggtgttaa gtcaagtagt ttaaggtact actctgtaag ataacacaga aaacagccaa
     9181 cctaaccgaa aagcgaaagc tgatacggga acagagcacg gttggaaagc gatgagttac
     9241 ctaaagacaa tcgggtacga ctgagtcgca atgttaatca gatataaggt ataagttgtg
     9301 tttactgaac gcaagtttct aatttcggtt aaatctcgat agaggaaagt gtctgaaacc
     9361 tctagtacaa agaaaggtaa gttagcattg atgacttatc tgttatcacc acatttgtac
     9421 aatctgtagg agaacctatg ggaacgaaac gaaagcgatg ccgagaatct gaatttacca
     9481 agacttaaca ctaactgggg ataccctaaa caagaatgcc taatagaaag gaggaaaaag
     9541 gctatagcac tagagcttga aaatcttgca agggtacgga gtactcgtag tagtctgaga
     9601 agggtaacgc cctttacatg gcaaaggggt acagttattg tgtactaaaa ttaaaaattg
     9661 attagggagg aaaacctcaa aatgaaacca acaatggcaa ttttagaaag aatcagtaaa
     9721 aattcacaag aaaatataga cgaagttttt acaagacttt atcgttatct tttacgtcca
     9781 gatatttatt acgtggcgac gcgtataact tcgtataatg tatgctatac gaacggtaac
     9841 gcgttgggaa atggcaatga tagcgaaaca acgtaaaact cttgttgtat gctttcattg
     9901 tcatcgtcac gtgattcata aacacaagtg aatgtcgact aatacgactc actatagggt
     9961 cgacagtgaa tttttacgaa cgaacaataa cagagccgta tactccgaga ggggtacgta
    10021 cggttcccga agagggtggt gcaaaccagt cacagtaatg tgaacaaggc ggtacctccc
    10081 tacttcacca tatcattttc tgcagccccc tagaaataat tttgtttaac tttaagaagg
    10141 agatatacat atatggctag atcgtccatt ccgacagcat cgccagtcac tatggcgtgc
    10201 tgctagcgct ata
//
