LOCUS       sequence-235310-        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(106..732)
                     /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     637..656
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    839..857
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     CDS             complement(2026..2397)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            2430..2539
                     /note="incP origin of transfer"
     primer_bind     complement(3730..3747)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3901..4489)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3981..4000)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 aatccttaca tctcccctaa tttttagact taagggcgaa ttctgcatca agctagcata
       61 aaaataagaa gcctgcattt gcaggcttct tatttttatg gtaccttaag acccactttc
      121 acatttaagt tgtttttcta atccgcatat gatcaattca aggccgaata agaaggctgg
      181 ctctgcacct tggtgatcaa ataattcgat agcttgtcgt aataatggcg gcatactatc
      241 agtagtaggt gtttcccttt cttctttagc gacttgatgc tcttgatctt ccaatacgca
      301 acctaaagta aaatgcccca cagcgctgag tgcatataat gcattctcta gtgaaaaacc
      361 ttgttggcat aaaaaggcta attgattttc gagagtttca tactgttttt ctgtaggccg
      421 tgtacctaaa tgtacttttg ctccatcgcg atgacttagt aaagcacatc taaaactttt
      481 agcgttatta cgtaaaaaat cttgccagct ttccccttct aaagggcaaa agtgagtatg
      541 gtgcctatct aacatctcaa tggctaaggc gtcgagcaaa gcccgcttat tttttacatg
      601 ccaatacaat gtaggctgct ctacacctag cttctgggcg agtttacggg ttgttaaacc
      661 ttcgattccg acctcattaa gcagctctaa tgcgctgtta atcactttac ttttatctaa
      721 tctagacatc attaattcct cctttttgtt gacattatat cattgataga gttatttgtc
      781 aaactagttt tttatttcga tgccctggac ttcatgaaaa actaaaaaaa atattgacac
      841 tctatcattg atagagtata attaaaataa gcttgatcgt agcgttaaca gatctgagct
      901 cctgcagtaa aggagaaaat tttatggttt caaaaggaga agaattattt acaggggtag
      961 taccaatttt agtagaactt gacggagatg taaatggaca caaattttca gtatcaggag
     1021 aaggagaagg agatgcaaca tacggaaaat taacacttaa attcatttgt acaacaggta
     1081 aactaccggt tccatggccc acacttgtga ctactttaac atggggagta caatgttttt
     1141 caagatatcc agatcatatg aaacaacacg acttttttaa atcagcaatg cctgaaggat
     1201 acgtacaaga aagaacaata ttttttaaag atgatggtaa ctataaaacg agagcagaag
     1261 taaaatttga aggagataca ttagtaaaca gaattgaatt aaaaggaata gactttaaag
     1321 aagatggaaa tatcctagga cataaattag aatacaatta catcagtcat aacgtataca
     1381 taacggcaga taaacagaaa aatggaatta aagctaactt taaaataaga cacaatatag
     1441 aagacggatc agtacaatta gcagatcact atcaacaaaa tacacctatc ggagatggcc
     1501 cagtgcttct accagataat cattacttaa gcacgcaaag cgctttatct aaagacccaa
     1561 atgaaaaaag agaccacatg gtattattag aattcgtaac agcagcagga attacactag
     1621 gaatggatga attatataag atgaacgcat gcgtcgactt aattaaacag ctgaggcgcg
     1681 ccggatccta taagttttaa taaaacttta aatagaaaaa ggcttctctc atgagaagtc
     1741 ttttttattt aaaataaata taaaataaaa tagaggctat aaatagcctc tattttatgt
     1801 gagaaatccc taaataaaaa gatgccagtg tgctggaatt cgcccttagg ctgcgctagg
     1861 gacctcttta gctccttgga agctgtcagt agtaggggat cggtcttgcc ttgctcgtcg
     1921 gtgatgtact tcaccagctc cgcgaagtcg ctcttcttga tggagcgcat ggggacgtgc
     1981 ttggcaatca cgcgcacccc ccggccgttt tagcggctaa aaaagtcatg gctctgccct
     2041 cgggcggacc acgcccatca tgaccttgcc aagctcgtcc tgcttctctt cgatcttcgc
     2101 cagcagggcg aggatcgtgg catcaccgaa ccgcgccgtg cgcgggtcgt cggtgagcca
     2161 gagtttcagc aggccgccca ggcggcccag gtcgccattg atgcgggcca gctcgcggac
     2221 gtgctcatag tccacgacgc ccgtgatttt gtagccctgg ccgacggcca gcaggtaggc
     2281 cgacaggctc atgccggccg ccgccgcctt ttcctcaatc gctcttcgtt cgtctggaag
     2341 gcagtacacc ttgataggtg ggctgccctt cctggttggc ttggtttcat cagccatccg
     2401 cttgccctca tctgttacgc cggcggtagc cggccagcct cgcagagcag gattcccgtt
     2461 gagcaccgcc aggtgcgaat aagggacagt gaagaaggaa cacccgctcg cgggtgggcc
     2521 tacttcacct atcctgcccg gctgacgccg ttggatacac caaggaaagt ctacacgaac
     2581 cctttggcaa aatcctgtat atcgtgcgaa aaaggatgga tataccgaaa aaatcgctat
     2641 aatgaccccg aagcaggaag ggcgaattct gcagatctga ccggtctctg aaaatataaa
     2701 aaccacagat tgatactaaa accttggttg tgttgctttt cggggcttaa atcaaggaaa
     2761 aatccttgtt ttaagccttt caaaaagaaa cacaaggtct ttgtactaac ctgtggttat
     2821 gtataaaatt gtagatttta gggtaacaaa aaacaccgta tttctacgat gtttttgctt
     2881 aaatacttgt ttttagttac agacaaacct gaagttaact atttatcaat tcctgcaatt
     2941 cgtttacaaa acggcaaatg tgaaatccgt cacatactgc gtgatgaact tgaattgcca
     3001 aaggaagtat aattttgtta tcttctttat aatatttccc catagtaaaa ataggaatca
     3061 aataatcata tcctttctgc aaattcagat taaagccatc gaaggttgac cacggtatca
     3121 tagatacatt aaaaatgttt tccggagcat ttggctttcc ttccattcta tgattgtttc
     3181 cataccgttg cgtatcactt tcataatctg ctaaaaatga tttaaagtca gacttacact
     3241 cagtccaaag gctggaaaat gtttcagtat cattgtgaaa tattgtatag cttggtatca
     3301 tctcatcata tatccccaat tcaccatctt gattgattgc cgtcctaaac tctgaatggc
     3361 ggtttacaat cattgcaata taataaagca ttgcaggata tagtttcatt cccttttcct
     3421 ttatttgtgt gatatccact ttaacggtca tgctgtaggt acaaggtaca cttgcaaagt
     3481 agtggtcaaa atactctttt ctgttccaac tatttttatc aattttttca aataccatct
     3541 aagttccctc tcaaattcaa gtttatcgct ctaatgaaca aagatattat accacatttt
     3601 tgtgaatttt tcaacttgcc cacttcgact gcactcccga cttaataact tcttgaacac
     3661 ttgccgaaaa agaaaaactg ccgggtacgt acccggggat cgatccccgg ccgagcgctc
     3721 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     3781 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     3841 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     3901 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     3961 gcgaaacccg acaggactat aaagatacca ggcgtttccc ccttgaagct ccctcgtgcg
     4021 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4081 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4141 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4201 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4261 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4321 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     4381 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4441 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     4501 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4561 catgaaatgc aagtttctaa ctaacataca tcatagttac taaactatgg taactataat
     4621 tttattaact atattaagca ctgattagta ctataactca atataagcat atcccctgta
     4681 tcgtaactag agaaccaaac gacggaaaaa gcgatataga taagtttaga tagtgcacca
     4741 attttttact tttctatttt agaatctagc atttccaatg cttactaata ctatttttga
     4801 tttttaactt gttttaaaaa agcacctact aattaggatt ctagctctat attctagtta
     4861 tacattctct atattctagc tctatattct agttatacat tctctatatt ctagctctat
     4921 attctagtta tacattctct atattctagc tctatattct agttatacat tctctatatt
     4981 ctagctctat attctagtta tacattctct atattctagc tctatattct agttatacat
     5041 tctctatatt ctagctctat attctagtta tacattctct aaaaaacata tatatatata
     5101 ctacatatat atgtacaatt attttaccaa aagaaatttt taaaacggct taatttcaat
     5161 actttgatgc catttttttt ttgcaattct cgtacaatta ttttaccaaa agaaattttt
     5221 attaattagt ttttttgcat aaaaatagga ctagaaaatc aattctaatc ctatcaaaaa
     5281 tctacttgtt taatctatat gtatagtaca taaatcctct atcatttatt ttagtttccc
     5341 atacatctaa aattttagta tctaataaag attctatcat atttgaaatt ctttgctgtg
     5401 ttactcctaa atcttttgct aattcctctt gattaattct taatatattt ccttttattt
     5461 tatcatcttt gaccaataaa tcttgcttat atctcatatg agtatataat cttaattctt
     5521 cagttgaaat tattccccaa attactgcca tggtaacaaa gtaactaata gtatattgtt
     5581 tatctatatt acatctaact tcttttacat tgtatgtatc ttttattcct agctttgtaa
     5641 caccttttgt ttttgtaata taatcattat gttgaagttc ttttaatgtt tctcttaatg
     5701 tcttttcact cataacaatg tttttaactt tcttcttacg tttataggtt atcagctcca
     5761 ttatatcgtc tatgtttaat tctctatctt tattgttaag taacacatta tagataaaca
     5821 attgattacc agtcattatt ttaacccctt tcctattctt atatttcaaa tcctttgagt
     5881 gcttatgtgg catattgaac aaagtatctt catcactata tataaaatct gattctattt
     5941 tattccaaca atcactatta aactcacaac ctgtacaacc aaaattaacc gttttaagct
     6001 tctcatacat atacttgact tgatattcta gctcactatt tgaaagacta ttttgtgata
     6061 tatagttcca ttctctagca acggatatta cttgacttaa acttttattc ctcaatctta
     6121 aagtcacaac tattttttgc aaatctaggt ttttatgtcc atctttacta ccatttttta
     6181 aaatatctac tatacaattt ggtagcttac tatctaatat atattttgta ttagtattac
     6241 ctactgtttt tacatctttg caatattttt tagctaactt ttctatgtca tatctatata
     6301 tatttttgtc ttggtgtatt atttttactt gtttagtagt atttttaata ttatatgtat
     6361 aaggtactct taatacttga gtagataaat ttgcttgttt atctgcaccc aatttattta
     6421 ttaatgtatt tgtaacttct gatactaact taatgttatt agttttatta attagcacat
     6481 aaacatggaa tccatttcca ctatcaacta ggatatgata atgtaatcct atcttagtaa
     6541 ataaattaat tatatcttta taattaaaac cttgtcctaa ttcttttttg tcaaaatcaa
     6601 atcctaatgc atatctattt attaagtttt cattggttcc tgcttttcca tcagttgtac
     6661 ataggttaaa aaatgtattt tcataataaa tattttttgc gaaggtaaat tctactatat
     6721 catctatatt tttaaaaaaa gttgttttac tatattcttt aaattcttct ccgagttcta
     6781 attgtttact attactttta ttattttgaa atactctaac atattcatta tcaataagat
     6841 tatttccttg tcctatacca tctctaaata ctgccattag aaattttttc aacttatatt
     6901 ttgaatctaa ttgttccata tatttattat ctccttatct agttttttta agagataaca
     6961 ctacattaaa tatatactta ttgactaatc aacttttaca actattgaat aattttagct
     7021 tagtatgtat aataatattg ttaagatgct aaaattgata gcaatattat tatataacta
     7081 cagatactta atcagttaga ccgatattgg agttggtagc tccataaact caattggtag
     7141 ttgagttgtc attgtctagt agtattatct cattttgttg ttatcttaaa atattatttt
     7201 aaaaatactc tgcttggtcg gcttagggta ttttttttgt ttaattccaa tattggttca
     7261 aactcctgtg taatactttt gacacttcca tctgtattga attgacagtt gatatggtat
     7321 ctcattattg gcttgttctt atgtttttct atgtatttat tttttacttt caaataattt
     7381 gtatttccat ttttagttat gtttaacaca ctatccttat tactcaataa aattattttt
     7441 tgttttaaca tatgatttat agaatctaaa aattgatatt gacttataat agtactttct
     7501 acctctctag ctttttctag tgtttcaaat tcttcaaact ctgctatatt gctatagtat
     7561 agcttatcat taataatatt atctgacaac tttaattcca ttttataaaa caattccaag
     7621 ttatgtataa aatttttaat gttacaacat atagacttat tatttgttct tatatggtat
     7681 ctccatttac tacttctagc tctcatgtta tatgcacaac tactcccaac ataaacatat
     7741 attgcttgtt ctaactcata tttttgtaaa atttcaacat catatccagt caaatcgtac
     7801 tctctcttat atacaaatgg tttttctaat tttttgttca ataaatctcc tcctatttct
     7861 tacttaattt agctaaaaat ctatttatat ttcacctcct ttttgacttt aagcctacga
     7921 ataccatatt tatgattata acatcttcta tttttccca
//
