LOCUS       sequence-229761-        7950 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli-C. difficile shuttle vector: tetracycline-inducible promoter
            (Ptet) driving expression of red fluorescent protein (mCherryOpt), 
            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 7950)
  AUTHORS   Ransom EM, Ellermeier CD, Weiss DS
  TITLE     Use of mCherry Red fluorescent protein for studies of protein 
            localization and gene expression in Clostridium difficile.
  JOURNAL   Appl Environ Microbiol. 2015 Mar;81(5):1652-60. doi: 
            10.1128/AEM.03446-14. Epub 2014 Dec 19.
  PUBMED    25527559
REFERENCE   2  (bases 1 to 7950)
  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/AEM.03446-14"; journalName: "Appl Environ Microbiol"; date:
            "2015-03"; volume: "81"; issue: "5"; pages: "1652-60"
COMMENT     Sequence Label: pRAN473
FEATURES             Location/Qualifiers
     source          1..7950
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(912..1538)
                     /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     1443..1462
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    1645..1663
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     CDS             complement(2823..3194)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            3227..3336
                     /note="incP origin of transfer"
     primer_bind     complement(4527..4544)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4698..5286)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4778..4797)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gtctagtagt attatctcat tttgttgtta tcttaaaata ttattttaaa aatactctgc
       61 ttggtcggct tagggtattt tttttgttta attccaatat tggttcaaac tcctgtgtaa
      121 tacttttgac acttccatct gtattgaatt gacagttgat atggtatctc attattggct
      181 tgttcttatg tttttctatg tatttatttt ttactttcaa ataatttgta tttccatttt
      241 tagttatgtt taacacacta tccttattac tcaataaaat tattttttgt tttaacatat
      301 gatttataga atctaaaaat tgatattgac ttataatagt actttctacc tctctagctt
      361 tttctagtgt ttcaaattct tcaaactctg ctatattgct atagtatagc ttatcattaa
      421 taatattatc tgacaacttt aattccattt tataaaacaa ttccaagtta tgtataaaat
      481 ttttaatgtt acaacatata gacttattat ttgttcttat atggtatctc catttactac
      541 ttctagctct catgttatat gcacaactac tcccaacata aacatatatt gcttgttcta
      601 actcatattt ttgtaaaatt tcaacatcat atccagtcaa atcgtactct ctcttatata
      661 caaatggttt ttctaatttt ttgttcaata aatctcctcc tatttcttac ttaatttagc
      721 taaaaatcta tttatatttc acctcctttt tgactttaag cctacgaata ccatatttat
      781 gattataaca tcttctattt ttcccaaatc cttacatctc ccctaatttt tagacttaag
      841 ggcgaattct gcatcaagct agcataaaaa taagaagcct gcatttgcag gcttcttatt
      901 tttatggtac cttaagaccc actttcacat ttaagttgtt tttctaatcc gcatatgatc
      961 aattcaaggc cgaataagaa ggctggctct gcaccttggt gatcaaataa ttcgatagct
     1021 tgtcgtaata atggcggcat actatcagta gtaggtgttt ccctttcttc tttagcgact
     1081 tgatgctctt gatcttccaa tacgcaacct aaagtaaaat gccccacagc gctgagtgca
     1141 tataatgcat tctctagtga aaaaccttgt tggcataaaa aggctaattg attttcgaga
     1201 gtttcatact gtttttctgt aggccgtgta cctaaatgta cttttgctcc atcgcgatga
     1261 cttagtaaag cacatctaaa acttttagcg ttattacgta aaaaatcttg ccagctttcc
     1321 ccttctaaag ggcaaaagtg agtatggtgc ctatctaaca tctcaatggc taaggcgtcg
     1381 agcaaagccc gcttattttt tacatgccaa tacaatgtag gctgctctac acctagcttc
     1441 tgggcgagtt tacgggttgt taaaccttcg attccgacct cattaagcag ctctaatgcg
     1501 ctgttaatca ctttactttt atctaatcta gacatcatta attcctcctt tttgttgaca
     1561 ttatatcatt gatagagtta tttgtcaaac tagtttttta tttcgatgcc ctggacttca
     1621 tgaaaaacta aaaaaaatat tgacactcta tcattgatag agtataatta aaataagctt
     1681 gatcgtagcg ttaacagatc tgagctcctg cagtaaagga gaaaatttta tggtatctaa
     1741 aggagaagaa gataatatgg ctataattaa agaatttatg agatttaaag ttcatatgga
     1801 aggatctgtt aatggacatg aatttgaaat agaaggagaa ggagaaggaa gaccttatga
     1861 aggtactcaa acagctaaat taaaagtaac taaaggtggt ccattaccat ttgcatggga
     1921 tatattaagt cctcaattta tgtatggatc taaagcatat gttaaacatc ctgctgatat
     1981 acctgattat ttaaaattat catttccaga aggttttaaa tgggaaagag ttatgaattt
     2041 tgaagatggt ggagtagtta ctgtaacaca agatagttca ttacaagatg gagattttat
     2101 atataaagtt aaattaagag gtactaattt tccatctgat ggaccagtaa tgcaaaagaa
     2161 aactatggga tgggaagcat catctgaaag aatgtatcca gaagatggag cattaaaagg
     2221 tgaaattaaa caaagattaa aattaaaaga tggaggtcat tatgatgctg aagttaagac
     2281 tacatataaa gctaagaaac cagttcaatt accaggtgca tataatgtta atattaaatt
     2341 agatataact tctcataatg aagattatac tatagttgaa caatatgaaa gagcagaagg
     2401 aagacattca acaggaggta tggatgaatt atataaaatg aacgcatgcg tcgacttaat
     2461 taaacagctg aggcgcgccg gatcctataa gttttaataa aactttaaat agaaaaaggc
     2521 ttctctcatg agaagtcttt tttatttaaa ataaatataa aataaaatag aggctataaa
     2581 tagcctctat tttatgtgag aaatccctaa ataaaaagat gccagtgtgc tggaattcgc
     2641 ccttaggctg cgctagggac ctctttagct ccttggaagc tgtcagtagt aggggatcgg
     2701 tcttgccttg ctcgtcggtg atgtacttca ccagctccgc gaagtcgctc ttcttgatgg
     2761 agcgcatggg gacgtgcttg gcaatcacgc gcaccccccg gccgttttag cggctaaaaa
     2821 agtcatggct ctgccctcgg gcggaccacg cccatcatga ccttgccaag ctcgtcctgc
     2881 ttctcttcga tcttcgccag cagggcgagg atcgtggcat caccgaaccg cgccgtgcgc
     2941 gggtcgtcgg tgagccagag tttcagcagg ccgcccaggc ggcccaggtc gccattgatg
     3001 cgggccagct cgcggacgtg ctcatagtcc acgacgcccg tgattttgta gccctggccg
     3061 acggccagca ggtaggccga caggctcatg ccggccgccg ccgccttttc ctcaatcgct
     3121 cttcgttcgt ctggaaggca gtacaccttg ataggtgggc tgcccttcct ggttggcttg
     3181 gtttcatcag ccatccgctt gccctcatct gttacgccgg cggtagccgg ccagcctcgc
     3241 agagcaggat tcccgttgag caccgccagg tgcgaataag ggacagtgaa gaaggaacac
     3301 ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct gacgccgttg gatacaccaa
     3361 ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc gtgcgaaaaa ggatggatat
     3421 accgaaaaaa tcgctataat gaccccgaag caggaagggc gaattctgca gatctgaccg
     3481 gtctctgaaa atataaaaac cacagattga tactaaaacc ttggttgtgt tgcttttcgg
     3541 ggcttaaatc aaggaaaaat ccttgtttta agcctttcaa aaagaaacac aaggtctttg
     3601 tactaacctg tggttatgta taaaattgta gattttaggg taacaaaaaa caccgtattt
     3661 ctacgatgtt tttgcttaaa tacttgtttt tagttacaga caaacctgaa gttaactatt
     3721 tatcaattcc tgcaattcgt ttacaaaacg gcaaatgtga aatccgtcac atactgcgtg
     3781 atgaacttga attgccaaag gaagtataat tttgttatct tctttataat atttccccat
     3841 agtaaaaata ggaatcaaat aatcatatcc tttctgcaaa ttcagattaa agccatcgaa
     3901 ggttgaccac ggtatcatag atacattaaa aatgttttcc ggagcatttg gctttccttc
     3961 cattctatga ttgtttccat accgttgcgt atcactttca taatctgcta aaaatgattt
     4021 aaagtcagac ttacactcag tccaaaggct ggaaaatgtt tcagtatcat tgtgaaatat
     4081 tgtatagctt ggtatcatct catcatatat ccccaattca ccatcttgat tgattgccgt
     4141 cctaaactct gaatggcggt ttacaatcat tgcaatataa taaagcattg caggatatag
     4201 tttcattccc ttttccttta tttgtgtgat atccacttta acggtcatgc tgtaggtaca
     4261 aggtacactt gcaaagtagt ggtcaaaata ctcttttctg ttccaactat ttttatcaat
     4321 tttttcaaat accatctaag ttccctctca aattcaagtt tatcgctcta atgaacaaag
     4381 atattatacc acatttttgt gaatttttca acttgcccac ttcgactgca ctcccgactt
     4441 aataacttct tgaacacttg ccgaaaaaga aaaactgccg ggtacgtacc cggggatcga
     4501 tccccggccg agcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     4561 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     4621 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     4681 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     4741 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     4801 tgaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     4861 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     4921 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     4981 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     5041 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     5101 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct
     5161 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     5221 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     5281 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     5341 cgttaaggga ttttggtcat gaaatgcaag tttctaacta acatacatca tagttactaa
     5401 actatggtaa ctataatttt attaactata ttaagcactg attagtacta taactcaata
     5461 taagcatatc ccctgtatcg taactagaga accaaacgac ggaaaaagcg atatagataa
     5521 gtttagatag tgcaccaatt ttttactttt ctattttaga atctagcatt tccaatgctt
     5581 actaatacta tttttgattt ttaacttgtt ttaaaaaagc acctactaat taggattcta
     5641 gctctatatt ctagttatac attctctata ttctagctct atattctagt tatacattct
     5701 ctatattcta gctctatatt ctagttatac attctctata ttctagctct atattctagt
     5761 tatacattct ctatattcta gctctatatt ctagttatac attctctata ttctagctct
     5821 atattctagt tatacattct ctatattcta gctctatatt ctagttatac attctctaaa
     5881 aaacatatat atatatacta catatatatg tacaattatt ttaccaaaag aaatttttaa
     5941 aacggcttaa tttcaatact ttgatgccat tttttttttg caattctcgt acaattattt
     6001 taccaaaaga aatttttatt aattagtttt tttgcataaa aataggacta gaaaatcaat
     6061 tctaatccta tcaaaaatct acttgtttaa tctatatgta tagtacataa atcctctatc
     6121 atttatttta gtttcccata catctaaaat tttagtatct aataaagatt ctatcatatt
     6181 tgaaattctt tgctgtgtta ctcctaaatc ttttgctaat tcctcttgat taattcttaa
     6241 tatatttcct tttattttat catctttgac caataaatct tgcttatatc tcatatgagt
     6301 atataatctt aattcttcag ttgaaattat tccccaaatt actgccatgg taacaaagta
     6361 actaatagta tattgtttat ctatattaca tctaacttct tttacattgt atgtatcttt
     6421 tattcctagc tttgtaacac cttttgtttt tgtaatataa tcattatgtt gaagttcttt
     6481 taatgtttct cttaatgtct tttcactcat aacaatgttt ttaactttct tcttacgttt
     6541 ataggttatc agctccatta tatcgtctat gtttaattct ctatctttat tgttaagtaa
     6601 cacattatag ataaacaatt gattaccagt cattatttta acccctttcc tattcttata
     6661 tttcaaatcc tttgagtgct tatgtggcat attgaacaaa gtatcttcat cactatatat
     6721 aaaatctgat tctattttat tccaacaatc actattaaac tcacaacctg tacaaccaaa
     6781 attaaccgtt ttaagcttct catacatata cttgacttga tattctagct cactatttga
     6841 aagactattt tgtgatatat agttccattc tctagcaacg gatattactt gacttaaact
     6901 tttattcctc aatcttaaag tcacaactat tttttgcaaa tctaggtttt tatgtccatc
     6961 tttactacca ttttttaaaa tatctactat acaatttggt agcttactat ctaatatata
     7021 ttttgtatta gtattaccta ctgtttttac atctttgcaa tattttttag ctaacttttc
     7081 tatgtcatat ctatatatat ttttgtcttg gtgtattatt tttacttgtt tagtagtatt
     7141 tttaatatta tatgtataag gtactcttaa tacttgagta gataaatttg cttgtttatc
     7201 tgcacccaat ttatttatta atgtatttgt aacttctgat actaacttaa tgttattagt
     7261 tttattaatt agcacataaa catggaatcc atttccacta tcaactagga tatgataatg
     7321 taatcctatc ttagtaaata aattaattat atctttataa ttaaaacctt gtcctaattc
     7381 ttttttgtca aaatcaaatc ctaatgcata tctatttatt aagttttcat tggttcctgc
     7441 ttttccatca gttgtacata ggttaaaaaa tgtattttca taataaatat tttttgcgaa
     7501 ggtaaattct actatatcat ctatattttt aaaaaaagtt gttttactat attctttaaa
     7561 ttcttctccg agttctaatt gtttactatt acttttatta ttttgaaata ctctaacata
     7621 ttcattatca ataagattat ttccttgtcc tataccatct ctaaatactg ccattagaaa
     7681 ttttttcaac ttatattttg aatctaattg ttccatatat ttattatctc cttatctagt
     7741 ttttttaaga gataacacta cattaaatat atacttattg actaatcaac ttttacaact
     7801 attgaataat tttagcttag tatgtataat aatattgtta agatgctaaa attgatagca
     7861 atattattat ataactacag atacttaatc agttagaccg atattggagt tggtagctcc
     7921 ataaactcaa ttggtagttg agttgtcatt
//
