LOCUS       sequence-229754-        7959 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli-C. difficile shuttle vector: tetracycline-inducible promoter
            (Ptet) driving expression of cyan fluorescent protein (CfpOpt), 
            multiple-cloning site for fusion to N-terminus of target protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7959)
  AUTHORS   Ransom EM, Williams KB, Weiss DS, Ellermeier CD
  TITLE     Identification and characterization of a gene cluster required for 
            proper rod shape, cell division, and pathogenesis in Clostridium 
            difficile.
  JOURNAL   J Bacteriol. 2014 Jun;196(12):2290-300. doi: 10.1128/JB.00038-14. 
            Epub 2014 Apr 11.
  PUBMED    24727226
REFERENCE   2  (bases 1 to 7959)
  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"; doi: 
            "10.1128/JB.00038-14"; journalName: "J Bacteriol"; date: "2014-06"; 
            volume: "196"; issue: "12"; pages: "2290-300"
COMMENT     Sequence Label: pRAN357
FEATURES             Location/Qualifiers
     source          1..7959
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(924..1550)
                     /codon_start=1
                     /gene="tetR"
                     /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="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     1455..1474
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    1657..1675
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     CDS             complement(2844..3215)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            3248..3357
                     /note="incP origin of transfer"
     primer_bind     complement(4548..4565)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4719..5307)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4799..4818)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgagttgtca ttgtctagta gtattatctc attttgttgt tatcttaaaa tattatttta
       61 aaaatactct gcttggtcgg cttagggtat tttttttgtt taattccaat attggttcaa
      121 actcctgtgt aatacttttg acacttccat ctgtattgaa ttgacagttg atatggtatc
      181 tcattattgg cttgttctta tgtttttcta tgtatttatt ttttactttc aaataatttg
      241 tatttccatt tttagttatg tttaacacac tatccttatt actcaataaa attatttttt
      301 gttttaacat atgatttata gaatctaaaa attgatattg acttataata gtactttcta
      361 cctctctagc tttttctagt gtttcaaatt cttcaaactc tgctatattg ctatagtata
      421 gcttatcatt aataatatta tctgacaact ttaattccat tttataaaac aattccaagt
      481 tatgtataaa atttttaatg ttacaacata tagacttatt atttgttctt atatggtatc
      541 tccatttact acttctagct ctcatgttat atgcacaact actcccaaca taaacatata
      601 ttgcttgttc taactcatat ttttgtaaaa tttcaacatc atatccagtc aaatcgtact
      661 ctctcttata tacaaatggt ttttctaatt ttttgttcaa taaatctcct cctatttctt
      721 acttaattta gctaaaaatc tatttatatt tcacctcctt tttgacttta agcctacgaa
      781 taccatattt atgattataa catcttctat ttttcccaaa tccttacatc tcccctaatt
      841 tttagactta agggcgaatt ctgcatcaag ctagcataaa aataagaagc ctgcatttgc
      901 aggcttctta tttttatggt accttaagac ccactttcac atttaagttg tttttctaat
      961 ccgcatatga tcaattcaag gccgaataag aaggctggct ctgcaccttg gtgatcaaat
     1021 aattcgatag cttgtcgtaa taatggcggc atactatcag tagtaggtgt ttccctttct
     1081 tctttagcga cttgatgctc ttgatcttcc aatacgcaac ctaaagtaaa atgccccaca
     1141 gcgctgagtg catataatgc attctctagt gaaaaacctt gttggcataa aaaggctaat
     1201 tgattttcga gagtttcata ctgtttttct gtaggccgtg tacctaaatg tacttttgct
     1261 ccatcgcgat gacttagtaa agcacatcta aaacttttag cgttattacg taaaaaatct
     1321 tgccagcttt ccccttctaa agggcaaaag tgagtatggt gcctatctaa catctcaatg
     1381 gctaaggcgt cgagcaaagc ccgcttattt tttacatgcc aatacaatgt aggctgctct
     1441 acacctagct tctgggcgag tttacgggtt gttaaacctt cgattccgac ctcattaagc
     1501 agctctaatg cgctgttaat cactttactt ttatctaatc tagacatcat taattcctcc
     1561 tttttgttga cattatatca ttgatagagt tatttgtcaa actagttttt tatttcgatg
     1621 ccctggactt catgaaaaac taaaaaaaat attgacactc tatcattgat agagtataat
     1681 taaaataagc ttgatcgtag cgttaacaga tctgagctcc tgcagtaaag gagaaaattt
     1741 tatggtttca aaaggagaag aattatttac aggggtagta ccaattttag tagaacttga
     1801 cggagatgta aatggacaca aattttcagt atcaggagaa ggagaaggag atgcaacata
     1861 cggaaaatta acacttaaat tcatttgtac aacaggtaaa ctaccggttc catggcccac
     1921 acttgtgact actttaacat ggggagtaca atgtttttca agatatccag atcatatgaa
     1981 acaacacgac ttttttaaat cagcaatgcc tgaaggatac gtacaagaaa gaacaatatt
     2041 ttttaaagat gatggtaact ataaaacgag agcagaagta aaatttgaag gagatacatt
     2101 agtaaacaga attgaattaa aaggaataga ctttaaagaa gatggaaata tcctaggaca
     2161 taaattagaa tacaattaca tcagtcataa cgtatacata acggcagata aacagaaaaa
     2221 tggaattaaa gctaacttta aaataagaca caatatagaa gacggatcag tacaattagc
     2281 agatcactat caacaaaata cacctatcgg agatggccca gtgcttctac cagataatca
     2341 ttacttaagc acgcaaagcg ctttatctaa agacccaaat gaaaaaagag accacatggt
     2401 attattagaa ttcgtaacag cagcaggaat tacactagga atggatgaat tatataagat
     2461 gaacgcatgc gtcgacttaa ttaaacagct gaggcgcgcc ggatcctata agttttaata
     2521 aaactttaaa tagaaaaagg cttctctcat gagaagtctt ttttatttaa aataaatata
     2581 aaataaaata gaggctataa atagcctcta ttttatgtga gaaatcccta aataaaaaga
     2641 tgccagtgtg ctggaattcg cccttaggct gcgctaggga cctctttagc tccttggaag
     2701 ctgtcagtag taggggatcg gtcttgcctt gctcgtcggt gatgtacttc accagctccg
     2761 cgaagtcgct cttcttgatg gagcgcatgg ggacgtgctt ggcaatcacg cgcacccccc
     2821 ggccgtttta gcggctaaaa aagtcatggc tctgccctcg ggcggaccac gcccatcatg
     2881 accttgccaa gctcgtcctg cttctcttcg atcttcgcca gcagggcgag gatcgtggca
     2941 tcaccgaacc gcgccgtgcg cgggtcgtcg gtgagccaga gtttcagcag gccgcccagg
     3001 cggcccaggt cgccattgat gcgggccagc tcgcggacgt gctcatagtc cacgacgccc
     3061 gtgattttgt agccctggcc gacggccagc aggtaggccg acaggctcat gccggccgcc
     3121 gccgcctttt cctcaatcgc tcttcgttcg tctggaaggc agtacacctt gataggtggg
     3181 ctgcccttcc tggttggctt ggtttcatca gccatccgct tgccctcatc tgttacgccg
     3241 gcggtagccg gccagcctcg cagagcagga ttcccgttga gcaccgccag gtgcgaataa
     3301 gggacagtga agaaggaaca cccgctcgcg ggtgggccta cttcacctat cctgcccggc
     3361 tgacgccgtt ggatacacca aggaaagtct acacgaaccc tttggcaaaa tcctgtatat
     3421 cgtgcgaaaa aggatggata taccgaaaaa atcgctataa tgaccccgaa gcaggaaggg
     3481 cgaattctgc agatctgacc ggtctctgaa aatataaaaa ccacagattg atactaaaac
     3541 cttggttgtg ttgcttttcg gggcttaaat caaggaaaaa tccttgtttt aagcctttca
     3601 aaaagaaaca caaggtcttt gtactaacct gtggttatgt ataaaattgt agattttagg
     3661 gtaacaaaaa acaccgtatt tctacgatgt ttttgcttaa atacttgttt ttagttacag
     3721 acaaacctga agttaactat ttatcaattc ctgcaattcg tttacaaaac ggcaaatgtg
     3781 aaatccgtca catactgcgt gatgaacttg aattgccaaa ggaagtataa ttttgttatc
     3841 ttctttataa tatttcccca tagtaaaaat aggaatcaaa taatcatatc ctttctgcaa
     3901 attcagatta aagccatcga aggttgacca cggtatcata gatacattaa aaatgttttc
     3961 cggagcattt ggctttcctt ccattctatg attgtttcca taccgttgcg tatcactttc
     4021 ataatctgct aaaaatgatt taaagtcaga cttacactca gtccaaaggc tggaaaatgt
     4081 ttcagtatca ttgtgaaata ttgtatagct tggtatcatc tcatcatata tccccaattc
     4141 accatcttga ttgattgccg tcctaaactc tgaatggcgg tttacaatca ttgcaatata
     4201 ataaagcatt gcaggatata gtttcattcc cttttccttt atttgtgtga tatccacttt
     4261 aacggtcatg ctgtaggtac aaggtacact tgcaaagtag tggtcaaaat actcttttct
     4321 gttccaacta tttttatcaa ttttttcaaa taccatctaa gttccctctc aaattcaagt
     4381 ttatcgctct aatgaacaaa gatattatac cacatttttg tgaatttttc aacttgccca
     4441 cttcgactgc actcccgact taataacttc ttgaacactt gccgaaaaag aaaaactgcc
     4501 gggtacgtac ccggggatcg atccccggcc gagcgctctt ccgcttcctc gctcactgac
     4561 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4621 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4681 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4741 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4801 agataccagg cgtttccccc ttgaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4861 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4921 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4981 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     5041 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5101 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     5161 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5221 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5281 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5341 gctcagtgga acgaaaactc acgttaaggg attttggtca tgaaatgcaa gtttctaact
     5401 aacatacatc atagttacta aactatggta actataattt tattaactat attaagcact
     5461 gattagtact ataactcaat ataagcatat cccctgtatc gtaactagag aaccaaacga
     5521 cggaaaaagc gatatagata agtttagata gtgcaccaat tttttacttt tctattttag
     5581 aatctagcat ttccaatgct tactaatact atttttgatt tttaacttgt tttaaaaaag
     5641 cacctactaa ttaggattct agctctatat tctagttata cattctctat attctagctc
     5701 tatattctag ttatacattc tctatattct agctctatat tctagttata cattctctat
     5761 attctagctc tatattctag ttatacattc tctatattct agctctatat tctagttata
     5821 cattctctat attctagctc tatattctag ttatacattc tctatattct agctctatat
     5881 tctagttata cattctctaa aaaacatata tatatatact acatatatat gtacaattat
     5941 tttaccaaaa gaaattttta aaacggctta atttcaatac tttgatgcca tttttttttt
     6001 gcaattctcg tacaattatt ttaccaaaag aaatttttat taattagttt ttttgcataa
     6061 aaataggact agaaaatcaa ttctaatcct atcaaaaatc tacttgttta atctatatgt
     6121 atagtacata aatcctctat catttatttt agtttcccat acatctaaaa ttttagtatc
     6181 taataaagat tctatcatat ttgaaattct ttgctgtgtt actcctaaat cttttgctaa
     6241 ttcctcttga ttaattctta atatatttcc ttttatttta tcatctttga ccaataaatc
     6301 ttgcttatat ctcatatgag tatataatct taattcttca gttgaaatta ttccccaaat
     6361 tactgccatg gtaacaaagt aactaatagt atattgttta tctatattac atctaacttc
     6421 ttttacattg tatgtatctt ttattcctag ctttgtaaca ccttttgttt ttgtaatata
     6481 atcattatgt tgaagttctt ttaatgtttc tcttaatgtc ttttcactca taacaatgtt
     6541 tttaactttc ttcttacgtt tataggttat cagctccatt atatcgtcta tgtttaattc
     6601 tctatcttta ttgttaagta acacattata gataaacaat tgattaccag tcattatttt
     6661 aacccctttc ctattcttat atttcaaatc ctttgagtgc ttatgtggca tattgaacaa
     6721 agtatcttca tcactatata taaaatctga ttctatttta ttccaacaat cactattaaa
     6781 ctcacaacct gtacaaccaa aattaaccgt tttaagcttc tcatacatat acttgacttg
     6841 atattctagc tcactatttg aaagactatt ttgtgatata tagttccatt ctctagcaac
     6901 ggatattact tgacttaaac ttttattcct caatcttaaa gtcacaacta ttttttgcaa
     6961 atctaggttt ttatgtccat ctttactacc attttttaaa atatctacta tacaatttgg
     7021 tagcttacta tctaatatat attttgtatt agtattacct actgttttta catctttgca
     7081 atatttttta gctaactttt ctatgtcata tctatatata tttttgtctt ggtgtattat
     7141 ttttacttgt ttagtagtat ttttaatatt atatgtataa ggtactctta atacttgagt
     7201 agataaattt gcttgtttat ctgcacccaa tttatttatt aatgtatttg taacttctga
     7261 tactaactta atgttattag ttttattaat tagcacataa acatggaatc catttccact
     7321 atcaactagg atatgataat gtaatcctat cttagtaaat aaattaatta tatctttata
     7381 attaaaacct tgtcctaatt cttttttgtc aaaatcaaat cctaatgcat atctatttat
     7441 taagttttca ttggttcctg cttttccatc agttgtacat aggttaaaaa atgtattttc
     7501 ataataaata ttttttgcga aggtaaattc tactatatca tctatatttt taaaaaaagt
     7561 tgttttacta tattctttaa attcttctcc gagttctaat tgtttactat tacttttatt
     7621 attttgaaat actctaacat attcattatc aataagatta tttccttgtc ctataccatc
     7681 tctaaatact gccattagaa attttttcaa cttatatttt gaatctaatt gttccatata
     7741 tttattatct ccttatctag tttttttaag agataacact acattaaata tatacttatt
     7801 gactaatcaa cttttacaac tattgaataa ttttagctta gtatgtataa taatattgtt
     7861 aagatgctaa aattgatagc aatattatta tataactaca gatacttaat cagttagacc
     7921 gatattggag ttggtagctc cataaactca attggtagt
//
