LOCUS       V003237                 8538 bp    DNA     circular SYN 18-DEC-2018
DEFINITION  Exported.
ACCESSION   V003237
VERSION     V003237
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8538)
  AUTHORS   Hon S, Holwerda EK, Worthen RS, Maloney MI, Tian L, Cui J, Lin PP,
            Lynd LR, Olson DG.
  TITLE     Expressing the Thermoanaerobacterium saccharolyticum pforA in
            engineered Clostridium thermocellum improves ethanol production
  JOURNAL   Biotechnol Biofuels 11, 242 (2018)
   PUBMED   30202437
REFERENCE   2  (bases 1 to 8538)
  AUTHORS   Hon S, Olson DG, Holwerda EK, Worthen RS, Maloney MI, Tian L, Cui J,
            Lin PP, Lynd LR.
  TITLE     Direct Submission
  JOURNAL   Submitted (20-APR-2018) Thayer School of Engineering, Dartmouth
            College, 14 Engineering Drive, Hanover, NH 03755, USA
REFERENCE   3  (bases 1 to 8538)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8538)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing
            ##Assembly-Data-END##
            SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol
            Biofuels 11, 242 (2018)"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (20-APR-2018) Thayer School of Engineering, Dartmouth College, 14
            Engineering Drive, Hanover, NH 03755, USA"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8538
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      28..573
                     /label="p15A ori"
                     /note="Plasmids containing the medium-copy-number p15A
                     origin of replication can be propagated in E. coli cells
                     that contain a second plasmid with the ColE1 origin."
     CDS             complement(840..1418)
                     /codon_start=1
                     /gene="tdk"
                     /product="thymidine kinase"
                     /label="tdk"
                     /protein_id="AYA22152.1"
                     /translation="MYGPKDHGYIEVVTGPMFSGKSEELIRRIKRAKIARQKVQVFKPA
                     IDDRYSIDKVVSHNGDNMHAIAIVKASDILAYAEEDTDVFAIDEVQFFDSEIVDIVKEI
                     ADSGKRVICAGLDMDFRGEPFGPTPELMAIAEFVDKLTAICMKCGNPATRTQRLINGKP
                     ANYDDPIIMVGAKESYEARCRKCHEVPRT"
     gene            complement(840..1418)
                     /gene="tdk"
                     /label="tdk"
     regulatory      complement(1419..2039)
                     /label="C. thermocellum cbp promoter"
                     /note="C. thermocellum cbp promoter"
                     /regulatory_class="promoter"
     CDS             complement(2118..3119)
                     /label="repB"
                     /note="RepB replication protein"
     rep_origin      complement(3120..3574)
                     /direction=LEFT
                     /label="C. thermocellum origin of replication"
                     /note="C. thermocellum origin of replication"
     CDS             complement(3680..4537)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(4538..4642)
                     /label="AmpR promoter"
     misc_feature    4732..5226
                     /label="upstream homology recombination flank"
                     /note="upstream homology recombination flank"
     misc_feature    5227..5777
                     /label="downstream homology recombination flank"
                     /note="downstream homology recombination flank"
     regulatory      5781..6355
                     /label="C. thermocellum gapDH promoter"
                     /note="C. thermocellum gapDH promoter"
                     /regulatory_class="promoter"
     CDS             6356..7003
                     /gene="cat"
                     /label="Chloramphenicol acetyltransferase"
                     /note="Chloramphenicol acetyltransferase from
                     Staphylococcus aureus. Accession#: P00485"
     CDS             7024..7569
                     /codon_start=1
                     /gene="hpt"
                     /product="hypoxanthine phosphoribosyltransferase"
                     /label="hpt"
                     /protein_id="AYA22156.1"
                     /translation="MENLSKDIDEILITEEELKEKIKELGRQITKDYKGKNLMLVGVLK
                     GALMFMADLSRHIDLPLSLDFMAVSSYGSSTHSSGIVKIIKDLDISIEGKDVLIVEDII
                     DSGLTLSYLRETLLGRKPKSLKICTILDKPERREASVKVDYVGFKIPDKFVVGYGLDFD
                     EKYRNLPFIGVLKPEMYS"
     gene            7024..7569
                     /gene="hpt"
                     /label="hpt"
     misc_feature    7754..8295
                     /label="gene target internal homology recombination flank"
                     /note="gene target internal homology recombination flank"
     terminator      8328..8516
                     /label="CYC1 terminator"
                     /note="transcription terminator for CYC1"
ORIGIN
        1 ttttatctga ttaataagat gatcttcttg agatcgtttt ggtctgcgcg taatctcttg
       61 ctctgaaaac gaaaaaaccg ccttgcaggg cggtttttcg aaggttctct gagctaccaa
      121 ctctttgaac cgaggtaact ggcttggagg agcgcagtca ccaaaacttg tcctttcagt
      181 ttagccttaa ccggcgcatg acttcaagac taactcctct aaatcaatta ccagtggctg
      241 ctgccagtgg tgcttttgca tgtctttccg ggttggactc aagacgatag ttaccggata
      301 aggcgcagcg gtcggactga acggggggtt cgtgcataca gtccagcttg gagcgaactg
      361 cctacccgga actgagtgtc aggcgtggaa tgagacaaac gcggccataa cagcggaatg
      421 acaccggtaa accgaaaggc aggaacagga gagcgcacga gggagccgcc agggggaaac
      481 gcctggtatc tttatagtcc tgtcgggttt cgccaccact gatttgagcg tcagatttcg
      541 tgatgcttgt caggggggcg gagcctatgg aaaaacggct ttgccgcggc cctctcactt
      601 ccctgttaag tatcttcctg gcatcttcca ggaaatctcc gccccgttcg taagccattt
      661 ccgctcgccg cagtcgaacg accgagcgta gcgagtcagt gagcgaggaa gcggaatata
      721 tcctgtatca catattctgc tgacgcaccg gtgcagcctt ttttctcctg ccacatgaag
      781 cacttcactg acaccctcat cagtgccaac atagtaagcc agtatacaaa aaaatccgct
      841 taagtccgcg ggacttcatg gcactttcta caccttgctt cataagactc ctttgctcca
      901 accattataa tgggatcgtc gtaattggca ggcttcccat ttataagcct ttgcgtgcga
      961 gtagcaggat tgccacactt catgcatata gcagtaagct tatcgacaaa ttcagctatg
     1021 gccatcaatt ctggagtcgg accaaatggt tcacctctaa agtccatgtc aagccctgca
     1081 catatgactc ttttcccgct atcggcaatc tcttttacga tgtcgactat ttcagaatca
     1141 aaaaattgaa cttcatctat agcaaataca tccgtatctt cttcagcata agccaatatg
     1201 tcagaagcct ttactatggc aatggcgtgc atgttgtcgc cgttatgaga tacgactttg
     1261 tctatggaat acctatcgtc tatagccggt ttgaaaacct gaactttttg cctggcaatc
     1321 ttagctctct ttatccttct tataagttcc tcactttttc cactgaacat gggaccagtt
     1381 acaacttcta tgtacccgtg gtctttaggc ccatacatct ttcgtcctcc ttaaaatttt
     1441 cgtttatttg ttttttattt agtgtgccta aaagcaccat cttatacatt cgtaccttat
     1501 atctcttatc ttgtatctct tacctcatat ctcttattat taactaccga agtatatttt
     1561 cataacggaa tcttatgact tatttttctg actgccgttt taccatacgg ttaatttata
     1621 cggaaacagc taatctaacc ttgctgaagg catgtttacc aaaatattat atctacttta
     1681 ccatattaat tcaagtggtc attttgatat tttacgccaa aatttttagc gttcctttac
     1741 ttgataattt atccaaaaac cttcattcgt caaaacaatt atgatataat taatttatga
     1801 tatgattaat taatatttac ggataactaa tcatttcaat gcaatagatc aacatgcaaa
     1861 tcaatatcca ccttactccg aacattttgc accaacgaat gcccagcact atatcccggc
     1921 aagcaatata acgtctataa ttttttatga taacgataaa attaaatttt atgaaagggg
     1981 taaaagcatg agaaaaatag ctgtattgtt aattactttg acgttcttag tcacgactcg
     2041 aatttaggag gcttacttgt ctgctttctt cattagaatc aatccttttt taaaagtcaa
     2101 tcccgtttgt tgaactactc tttaataaaa taatttttcc gttcccaatt ccacattgca
     2161 ataatagaaa atccatcttc atcggctttt tcgtcatcat ctgtatgaat caaatcgcct
     2221 tcttctgtgt catcaaggtt taatttttta tgtatttctt ttaacaaacc accataggag
     2281 attaaccttt tacggtgtaa accttcctcc aaatcagaca aacgtttcaa attcttttct
     2341 tcatcatcgg tcataaaatc cgtatccttt acaggatatt ttgcagtttc gtcaattgcc
     2401 gattgtatat ccgatttata tttatttttc ggtcgaatca tttgaacttt tacatttgga
     2461 tcatagtcta atttcattgc ctttttccaa aattgaatcc attgtttttg attcacgtag
     2521 ttttctgtat tcttaaaata agttggttcc acacatacca atacatgcat gtgctgatta
     2581 taagaattat ctttattatt tattgtcact tccgttgcac gcataaaacc aacaagattt
     2641 ttattaattt ttttatattg catcattcgg cgaaatcctt gagccatatc tgacaaactc
     2701 ttatttaatt cttcgccatc ataaacattt ttaactgtta atgtgagaaa caaccaacga
     2761 actgttggct tttgtttaat aacttcagca acaacctttt gtgactgaat gccatgtttc
     2821 attgctctcc tccagttgca cattggacaa agcctggatt tacaaaacca cactcgatac
     2881 aactttcttt cgcctgtttc acgattttgt ttatactcta atatttcagc acaatctttt
     2941 actctttcag cctttttaaa ttcaagaata tgcagaagtt caaagtaatc aacattagcg
     3001 attttctttt ctctccatgg tctcactttt ccactttttg tcttgtccac taaaaccctt
     3061 gatttttcat ctgaataaat gctactatta ggacacataa tattaaaaga aacccccatc
     3121 tatttagtta tttgtttggt cacttataac tttaacagat ggggtttttc tgtgcaacca
     3181 attttaaggg ttttcaatac tttaaaacac atacatacca acacttcaac gcacctttca
     3241 gcaactaaaa taaaaatgac gttatttcta tatgtatcaa gaatagaaag aactcgtttt
     3301 tcgctacgct caaaacgcaa aaaaagcact cattcgagtg ctttttctta tcgctccaaa
     3361 tcatgcgatt ttttcctctt tgcttttctt tgctcacgaa gttctcgatc acgctgcaaa
     3421 acatcttgaa gcgaaaaagt attcttcttt tcttccgatc gctcatgctg acgcacgaaa
     3481 agccctctag gcgcatagga acaactccta aatgcatgtg aggggttttc tcgtccatgt
     3541 gaacagtcgc atacgcaata ttttgtttcc catagcggcc gcgattatca aaaaggatct
     3601 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     3661 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     3721 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagc
     3781 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3841 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3901 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3961 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     4021 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     4081 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     4141 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     4201 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     4261 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     4321 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     4381 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     4441 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     4501 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     4561 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     4621 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     4681 ccattattat catgacatta acctataaaa ataggcgtat cacgaggcga ttgggttctt
     4741 atgctccgtg ggaggttcaa tgatttttgg tttaaaaaat gcaccgaatg tgggagcggc
     4801 agccatgctc atatctctga tagtggtacc tgtggtaagc ctctttacgg caaagcttcc
     4861 aaatgcgcat attaacatga tatacggtga ggaagacagt aagcagaaat tggcctcata
     4921 aaaggaaata tcttaacata aaatagaatt aacgtaaaat agaatttaat ttgtatattg
     4981 aaaaagcagc cgaagtttac ttttcggctg ctttgctttt tatggcgggg caatggatct
     5041 attttccgat tttaaaggtt ttcacttttg gactgcaggc tgtttacagc cttttttgaa
     5101 agcaattaat aattgacaat aatccgtcaa taaaagataa taataaaagg gtttaatatg
     5161 cttgaattat gtgataagat agaaataata ttattccatt atttatggtt agatggggga
     5221 ttgttgtata aatattgccg gcaaggcgga tatatcagga cagatatata aagtccacgg
     5281 gcgaagtgcc caaggtaata atataaaatt ataattttgt attgggggtt gtcgtatgaa
     5341 gtactggaat ccttcatgtg aatgtatgtc aagagatgaa atgacgaagc ttcagacgga
     5401 aaggctgatt gagactgtaa aaagagttta tcataatgtt ccgttgtata gggagaaaat
     5461 gcaaaagaag ggcattgagc ccggagacat taaaacgttg gatgacctga agaaacttcc
     5521 ttttacttac aaagacgatt tgagagatac atatccttat ggtatgtttg ccgctccgct
     5581 gagcgaaatt gtaagaatac acgcttcttc cggaacaacc ggaaaaccga cggtggtagg
     5641 ttacacgagg aaagacattg acacttggtc ggaggttacg gcaagaactt tggtttcagc
     5701 cggtgcagac aaaaattctc ttattcaggt tgcctatgga tacggacttt ttacgggagg
     5761 actgggaata cactacgatc gtgggaatag gcatggaaaa gatttttttg cccgaggatg
     5821 cattgataaa gtatttcttt tccgaaagag aggaaaagat tctaaagagt tttggaaata
     5881 ctgatgaata ttgtgtgcag agtacaattc tatggacaag aaaagaggct ttgtcaaaac
     5941 tttttcgtct gggaatgagg atggatttta aaaagctgga tactttggag ggcgaggtgg
     6001 tttttcagga aacaaacagg gcgcgtctgt tttcttttat atgcaataat tactgtatct
     6061 ctctggcatt gccaggtttt aataaagatt aaaattattg actagaaata aaaaaattgt
     6121 ccataatatt aatggacaaa aaaacaaaga attacatcaa aggaagataa aaatactttg
     6181 ttaaaaaatt aattattttt tatctaaact attgaaaatg aaaataaaat aatataaaat
     6241 gaatcatagt gcaagagata cttgccagag gatgaatatt ttactgcatt catgctttat
     6301 ggcagctaat agaggcatta aattaaattt taatttacaa taggaggcga tattaatgaa
     6361 ctttaataaa attgatttag acaattggaa gagaaaagag atatttaatc attatttgaa
     6421 ccaacaaacg acttttagta taaccacaga aattgatatt agtgttttat accgaaacat
     6481 aaaacaagaa ggatataaat tttaccctgc atttattttc ttagtgacaa gggtgataaa
     6541 ctcaaataca gcttttagaa ctggttacaa tagcgacgga gagttaggtt attgggataa
     6601 gttagagcca ctttatacaa tttttgatgg tgtatctaaa acattctctg gtatttggac
     6661 tcctgtaaag aatgacttca aagagtttta tgatttatac ctttctgatg tagagaaata
     6721 taatggttcg gggaaattgt ttcccaaaac acctatacct gaaaatgctt tttctctttc
     6781 tattattcca tggacttcat ttactgggtt taacttaaat atcaataata atagtaatta
     6841 ccttctaccc attattacag caggaaaatt cattaataaa ggtaattcaa tatatttacc
     6901 gctatcttta caggtacatc attctgtttg tgatggttat catgcaggat tgtttatgaa
     6961 ctctattcag gaattgtcag ataggcctaa tgactggctt ttataataaa ggaggtcgac
     7021 gtcatggaaa atttatcaaa agacatcgat gaaattttga tcacagaaga agaacttaag
     7081 gaaaagataa aagagcttgg gaggcaaatc acaaaagact acaaagggaa aaatttgatg
     7141 ttggtaggag ttttaaaagg tgctttaatg tttatggctg atttgtcaag acacatagat
     7201 ttgcctttat cacttgattt tatggctgtt tccagctatg gaagctcaac tcattcatca
     7261 ggaatagtaa agataatcaa agatcttgat ataagcatag aaggcaaaga tgttctgatt
     7321 gtggaagaca taattgacag cggtttgact ttgtcttact taagggaaac tttacttgga
     7381 aggaagccaa aaagcctgaa aatatgcaca atattagaca aaccggagag aagagaagca
     7441 tctgtaaaag tcgattatgt aggatttaag atacctgata agtttgtcgt gggttatgga
     7501 ttggactttg atgaaaagta caggaacctt ccttttatag gcgttttgaa acctgaaatg
     7561 tacagctaag cccgggcctc gagaaaacaa aaggctcagt cggaagactg ggccttttgt
     7621 tttggtaccg aattcggcgc gcctcagcgt ttaaacccgc tgatcctaga gggccgcatc
     7681 atgtaattag ttatgtcacg cttacattca cgccctcccc atgctttaat acatacctgg
     7741 cccagtagtt caggaatgta tgcaaaagct gcaaaagtgc ccatggttga gcctgccgac
     7801 agtagagagt gtaaagaata tgtgaaggcg gcttttgaaa taagcgaaaa atttgatact
     7861 ccggttattt tcagactttc gactaggata tcccattcac agagtgttgt tgaaatcggt
     7921 gaaagagagg atgtaccgct taaggaatac aaaaaggatc ccgaaaaata tgtcatgatg
     7981 cctgcaatgg cgcgtaaaag acatgttgcg gtagaaagca gaatggctgc tttggccgag
     8041 ttctccaata caactcctct aaacaggatt gaatggggaa gcacggatat cggtgtaata
     8101 accagcggaa ttgcctacca gtatgcaaga gaagccttcg gtgatgtctc atatctcaag
     8161 ctcggaatga tttatccact tccggacaag ttaattaaag agttttccga aaaagtaaaa
     8221 gtattatatg ttatagagga actggaaccg ttttttgaag agcatataag aaaactggga
     8281 atcaaggtta taggcctgct aatagtagtg aaaaaatcag ggataatgcc gctctaaccg
     8341 aaaaggaagg agttagacaa cctgaagtct aggtccctat ttattttttt atagttatgt
     8401 tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac agacgcgtgt
     8461 acgcatgtaa cattatactg aaaaccttgc ttgagaaggt tttgggacgc tcgaaggctt
     8521 taatttgcaa gctgcggc
//
