LOCUS       sequence-392933-        4772 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Replicating plasmid with homology region for introducing PolCC669Y 
            mutation. Confers thiamphenicol resistance..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4772)
  AUTHORS   Lanahan A, Zakowicz K, Tian L, Olson DG, Lynd LR
  TITLE     A Single Nucleotide Change in the polC DNA Polymerase III in 
            Clostridium thermocellum Is Sufficient To Create a Hypermutator 
            Phenotype.
  JOURNAL   Appl Environ Microbiol. 2022 Jan 11;88(1):e0153121. doi: 
            10.1128/AEM.01531-21. Epub 2021 Oct 20.
  PUBMED    35015978
REFERENCE   2  (bases 1 to 4772)
  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: "Appl 
            Environ Microbiol."; date: "2022-01-11"; volume: "88(1):e0153121. 
            doi"; pages: " 10.1128/AEM.01531-21. Epub 2021 Oct 20"
COMMENT     Sequence Label: pLT237
FEATURES             Location/Qualifiers
     source          1..4772
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..46)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(200..788)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(280..299)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(1071..1087)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1095..1111
                     /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        complement(join(1119..1125,1126..1143,1144..1149))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1164..1185
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3206..3222)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3230..3246
                     /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)."
     CDS             complement(3276..4280)
                     /codon_start=1
                     /gene="repB"
                     /product="RepB replication protein"
                     /label=repB
                     /note="from Enterococcus faecalis plasmid pAM-alpha-1"
                     /db_xref="GI:22652809"
                     /protein_id="AAN03827.1"
                     /translation="MGVSFNIMCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDY
                     FELLHILEFKKAERVKDCAEILEYKQNRETVERKLYRVWFCKSRLCPMCNWRRAMKHGI
                     QSQKVVAEVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLV
                     GFMRATEVTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPN
                     VKVQMIRPKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLIS
                     YGGLLKEIHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
ORIGIN
        1 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
       61 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
      121 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
      181 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
      241 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
      301 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
      361 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
      421 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
      481 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
      541 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
      601 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
      661 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
      721 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
      781 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
      841 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
      901 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
      961 accaatgctt aatcagtgag gcacctatct cagcgatctg tcaattcgag ctcggtaccc
     1021 ggggatcctc tagagtcgac ctgcaggcat gcaagcttgg cgtaatcatg gtcatagctg
     1081 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     1141 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     1201 ctgcccgctt tccagtcggg aaacctgtcg tgccagccct tcaaacttcc caaaggcgag
     1261 ccctagtgac attagaaaac cgactgtaaa aagtacagtc ggcattatct catattataa
     1321 aagccagtca ttaggcctat ctgacaattc ctgaatagag ttcataaaca atcctgcatg
     1381 ataaccatca caaacagaat gatgtacctg taaagatagc ggtaaatata ttgaattacc
     1441 tttattaatg aattttcctg ctgtaataat gggtagaagg taattactat tattattgat
     1501 atttaagtta aacccagtaa atgaagtcca tggaataata gaaagagaaa aagcattttc
     1561 aggtataggt gttttgggaa acaatttccc cgaaccatta tatttctcta catcagaaag
     1621 gtataaatca taaaactctt tgaagtcatt ctttacagga gtccaaatac cagagaatgt
     1681 tttagataca ccatcaaaaa ttgtataaag tggctctaac ttatcccaat aacctaactc
     1741 tccgtcgcta ttgtaaccag ttctaaaagc tgtatttgag tttatcaccc ttgtcactaa
     1801 gaaaataaat gcagggtaaa atttatatcc ttcttgtttt atgtttcggt ataaaacact
     1861 aatatcaatt tctgtggtta tactaaaagt cgtttgttgg ttcaaataat gattaaatat
     1921 ctcttttctc ttccaattgt ctaaatcaat tttattaaag ttcatttgat atgcctccta
     1981 aatttttatc taaagtgaat ttaggaggct tacttgtctg ctttcttcat tagaatcaat
     2041 ccttttttaa aagtcaatcc cgtttgtaaa gttaaacaaa attatttcta ttataggcaa
     2101 caaaggatga tccgacggat cccctcgacc ctacaagata gccgtcactt aaagatttcc
     2161 atacaagttt ttgcgcaata atatacataa cggagaagcc attttttatt attgagttaa
     2221 gctccttttc catgcgcttt tccacaatct caggaagagg gtcgccataa atttctctcg
     2281 ccttgttttc ggcaagcatc cttatttctt cttccgcacc ttcaatttta ggaggaaaag
     2341 ttccttccgg tatgggaagt atgtcctcgc acatatccgc aattaaattt gtatttgtta
     2401 caaccacttc atagcatttt tccctgccca gatattcaaa ctcttccagc atttcatcag
     2461 tggtcctgaa atacaacggt gcctggttgt ccgcatcgga aaaaccttgc cccgccataa
     2521 gtattcttct gaaaacttcg tcctgcggat ccatgaaatg aacatcgcag gtagcgacca
     2581 ccggcttgtt gtacttctca cccaaagcta ctatttttct gtttatattc tttaagtctt
     2641 cctcactgtt cacctttccg ctctcgacaa ggaaacggtt gttgcccaaa ggctgtattt
     2701 caagataatc gtaaaacttt acaatccttg aaatttcgtc ctcatctttg ttttcaagaa
     2761 tcgcacggta aagctctcca gcttcatatg cacttcccaa aatgagtcct tctctgtgca
     2821 tcattaaaag ctttcttgga agtctcggcc ttttgtgaaa atactccaaa tgagtcttgg
     2881 acacaatctt gtacagattt ctaaggccta cgtaattttt tacaagtata atggcatggt
     2941 atgaatttgc tttctgataa ttccaacaac cctcaaaaac attttgtatg tcatcaattg
     3001 tttttacatt cttttcagcc agtatctcaa ggcatttgat aaagatctcc ccacaggcct
     3061 ttgcatcgtc aagagccctg tgatggtttt ccaatgaaac accaagatgc tttgcaacca
     3121 catccagttt gtgtttttta agctccggaa acatttgcct tgaaagttcc aaagtgtcaa
     3181 ttacaggatt gtcggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     3241 tcacaattcc acacaacata cgagggatcc ttgaactact ctttaataaa ataatttttc
     3301 cgttcccaat tccacattgc aataatagaa aatccatctt catcggcttt ttcgtcatca
     3361 tctgtatgaa tcaaatcgcc ttcttctgtg tcatcaaggt ttaatttttt atgtatttct
     3421 tttaacaaac caccatagga gattaacctt ttacggtgta aaccttcctc caaatcagac
     3481 aaacgtttca aattcttttc ttcatcatcg gtcataaaat ccgtatcctt tacaggatat
     3541 tttgcagttt cgtcaattgc cgattgtata tccgatttat atttattttt cggtcgaatc
     3601 atttgaactt ttacatttgg atcatagtct aatttcattg cctttttcca aaattgaatc
     3661 cattgttttt gattcacgta gttttctgta ttcttaaaat aagttggttc cacacatacc
     3721 aatacatgca tgtgctgatt ataagaatta tctttattat ttattgtcac ttccgttgca
     3781 cgcataaaac caacaagatt tttattaatt tttttatatt gcatcattcg gcgaaatcct
     3841 tgagccatat ctgacaaact cttatttaat tcttcgccat cataaacatt tttaactgtt
     3901 aatgtgagaa acaaccaacg aactgttggc ttttgtttaa taacttcagc aacaaccttt
     3961 tgtgactgaa tgccatgttt cattgctctc ctccagttgc acattggaca aagcctggat
     4021 ttacaaaacc acactcgata caactttctt tcgactgttt cacgattttg tttatactct
     4081 aatatttcag cacaatcttt tactctttca gcctttttaa attcaagaat atgcagaagt
     4141 tcaaagtaat caacattagc gattttcttt tctctccatg gtctcacttt tccacttttt
     4201 gtcttgtcca ctaaaaccct tgatttttca tctgaataaa tgctactatt aggacacata
     4261 atattaaaag aaacccccat ctatttagtt atttgtttgg tcacttataa ctttaacaga
     4321 tggggttttt ctgtgcaacc aattttaagg gttttcaata ctttaaaaca catacatacc
     4381 aacacttcaa cgcacctttc agcaactaaa ataaaaatga cgttatttct atatgtatca
     4441 agaatagaaa gaactcgttt ttcgctacgc tcaaaacgca aaaaaagcac tcattcgagt
     4501 gctttttctt atcgctccaa atcatgcgat tttttcctct ttgcttttct ttgctcacga
     4561 agttctcgat cacgctgcaa aacatcttga agcgaaaaag tattcttctt ttcttccgat
     4621 cgctcatgct gacgcacgaa aagccctcta ggcgcatagg aacaactcct aaatgcatgt
     4681 gaggggtttt ctcgtccatg tgaacagtcg catacgcaat attttgtttc ccatactgca
     4741 ttaatgaatc ggccaacgcg cggggagagg cg
//
