LOCUS       V003240                 8488 bp    DNA     circular SYN 18-DEC-2018
DEFINITION  Exported.
ACCESSION   V003240
VERSION     V003240
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8488)
  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 8488)
  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 8488)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8488)
  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..8488
                     /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="AYA22157.1"
                     /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..5218
                     /label="upstream homology recombination flank"
                     /note="upstream homology recombination flank"
     misc_feature    5219..5750
                     /label="downstream homology recombination flank"
                     /note="downstream homology recombination flank"
     regulatory      5754..6328
                     /label="C. thermocellum gapDH promoter"
                     /note="C. thermocellum gapDH promoter"
                     /regulatory_class="promoter"
     CDS             6329..6976
                     /gene="cat"
                     /label="Chloramphenicol acetyltransferase"
                     /note="Chloramphenicol acetyltransferase from
                     Staphylococcus aureus. Accession#: P00485"
     CDS             6997..7542
                     /codon_start=1
                     /gene="hpt"
                     /product="hypoxanthine phosphoribosyltransferase"
                     /label="hpt"
                     /protein_id="AYA22161.1"
                     /translation="MENLSKDIDEILITEEELKEKIKELGRQITKDYKGKNLMLVGVLK
                     GALMFMADLSRHIDLPLSLDFMAVSSYGSSTHSSGIVKIIKDLDISIEGKDVLIVEDII
                     DSGLTLSYLRETLLGRKPKSLKICTILDKPERREASVKVDYVGFKIPDKFVVGYGLDFD
                     EKYRNLPFIGVLKPEMYS"
     gene            6997..7542
                     /gene="hpt"
                     /label="hpt"
     misc_feature    7727..8245
                     /label="gene target internal homology recombination flank"
                     /note="gene target internal homology recombination flank"
     terminator      8278..8466
                     /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 taataaagct
     4741 tgaagaaccg cagagtatta ccatcaagat tcatgatgtc gaaagatttg caaacagacg
     4801 tgacaactac agatacgata tatatttgtg ctcagtcata aaggcaaaca gaaatgacga
     4861 aaaaggcgtg tttgccatca tgactaatct aagccagggc ggggctgctt ttattgccaa
     4921 ggaagatatc gaaaaagaac tggaaacggc ttataatatg caggaggaga aaaattatta
     4981 ttttgaagta tttattactc ctaaaaaaca actgtgtttt accggtaccg tgaaaagaaa
     5041 aacgagtaat gaaaaaggat ttgagtatgg tgtaaaaatt ttcgatattg attttcaaaa
     5101 tgaaaaaact ctgaacgagt tgttggatga gcttgaaaaa aaagataaag agttctataa
     5161 taaacgaagt agcttttggt cgaagaatag caaatataat aaataatggg ggatttactg
     5221 cttaattaat gctacaatta tttgatatta agcattatta attaatatta aacattatta
     5281 aattcgaaat acggcttttt tgcattttaa tgtgaaatca ttgataaagc cctccttatt
     5341 gagaggaggg cctttttttg attgttttga tttttaattc ttatttttaa ttcttatacg
     5401 gtaaagatta ttacacgtta aagagaaggt cgccgtatgt cgggaacggc cacatttttg
     5461 aatcaaccag tgtttcgagt ttgtcagcca cggctctgag ttcattcatt ctgacgaaca
     5521 catcatccct gtacatacat gcttttttgt atgtgtcacc gctgaaatct gctgcctctg
     5581 aagtcgattt ttcaagagcg gccaagttct ttttaagttc ggcaagaagt gaggacactt
     5641 cctgcaaaag ctctgcctga acactggtgt cggcattgac acctgttgct ttgacagtgt
     5701 ttattgagtc ggcaagtttt gtaatatagc cgataacagc aggtataata atcgtgggaa
     5761 taggcatgga aaagattttt ttgcccgagg atgcattgat aaagtatttc ttttccgaaa
     5821 gagaggaaaa gattctaaag agttttggaa atactgatga atattgtgtg cagagtacaa
     5881 ttctatggac aagaaaagag gctttgtcaa aactttttcg tctgggaatg aggatggatt
     5941 ttaaaaagct ggatactttg gagggcgagg tggtttttca ggaaacaaac agggcgcgtc
     6001 tgttttcttt tatatgcaat aattactgta tctctctggc attgccaggt tttaataaag
     6061 attaaaatta ttgactagaa ataaaaaaat tgtccataat attaatggac aaaaaaacaa
     6121 agaattacat caaaggaaga taaaaatact ttgttaaaaa attaattatt ttttatctaa
     6181 actattgaaa atgaaaataa aataatataa aatgaatcat agtgcaagag atacttgcca
     6241 gaggatgaat attttactgc attcatgctt tatggcagct aatagaggca ttaaattaaa
     6301 ttttaattta caataggagg cgatattaat gaactttaat aaaattgatt tagacaattg
     6361 gaagagaaaa gagatattta atcattattt gaaccaacaa acgactttta gtataaccac
     6421 agaaattgat attagtgttt tataccgaaa cataaaacaa gaaggatata aattttaccc
     6481 tgcatttatt ttcttagtga caagggtgat aaactcaaat acagctttta gaactggtta
     6541 caatagcgac ggagagttag gttattggga taagttagag ccactttata caatttttga
     6601 tggtgtatct aaaacattct ctggtatttg gactcctgta aagaatgact tcaaagagtt
     6661 ttatgattta tacctttctg atgtagagaa atataatggt tcggggaaat tgtttcccaa
     6721 aacacctata cctgaaaatg ctttttctct ttctattatt ccatggactt catttactgg
     6781 gtttaactta aatatcaata ataatagtaa ttaccttcta cccattatta cagcaggaaa
     6841 attcattaat aaaggtaatt caatatattt accgctatct ttacaggtac atcattctgt
     6901 ttgtgatggt tatcatgcag gattgtttat gaactctatt caggaattgt cagataggcc
     6961 taatgactgg cttttataat aaaggaggtc gacgtcatgg aaaatttatc aaaagacatc
     7021 gatgaaattt tgatcacaga agaagaactt aaggaaaaga taaaagagct tgggaggcaa
     7081 atcacaaaag actacaaagg gaaaaatttg atgttggtag gagttttaaa aggtgcttta
     7141 atgtttatgg ctgatttgtc aagacacata gatttgcctt tatcacttga ttttatggct
     7201 gtttccagct atggaagctc aactcattca tcaggaatag taaagataat caaagatctt
     7261 gatataagca tagaaggcaa agatgttctg attgtggaag acataattga cagcggtttg
     7321 actttgtctt acttaaggga aactttactt ggaaggaagc caaaaagcct gaaaatatgc
     7381 acaatattag acaaaccgga gagaagagaa gcatctgtaa aagtcgatta tgtaggattt
     7441 aagatacctg ataagtttgt cgtgggttat ggattggact ttgatgaaaa gtacaggaac
     7501 cttcctttta taggcgtttt gaaacctgaa atgtacagct aagcccgggc ctcgagaaaa
     7561 caaaaggctc agtcggaaga ctgggccttt tgttttggta ccgaattcgg cgcgcctcag
     7621 cgtttaaacc cgctgatcct agagggccgc atcatgtaat tagttatgtc acgcttacat
     7681 tcacgccctc cccatgcttt aatacatacc tggcccagta gttcagttca tagacttata
     7741 gccgaggtta tagatgagct tgatattgaa ggaaagacaa taggtgtttc acctgttggc
     7801 tgtgcctaca acaactatga gtattttaat tgtgatatga ttcaggcagc ccatggaagg
     7861 gctccggcgg ttgcgaccgg ggcaaaaagg gttaatcccg acaatgttgt gttcgcctat
     7921 cagggggacg gagatttggc cgcgataggt acggcagaga ttgtccatgc agctacaagg
     7981 ggcgaaaaaa taacggttat ttttataaac aatgccattt acggaatgac atcgggacag
     8041 atggctccca ccaccttgat tgatcaggtt acgaccacaa ctccttttgg aagaaagccg
     8101 gaggttcacg gatatccgat aaatgtttgt gaacttttgt ccacacttga gggcgcggta
     8161 tatgttgaga gagtttcggt gcatgacata aaaaatatta tgaatgcaaa gaaggcaata
     8221 aagaaggcat ttaaagttca gatggctgct aatagtagtg aaaaaatcag ggataatgcc
     8281 gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat ttattttttt
     8341 atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac
     8401 agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt tttgggacgc
     8461 tcgaaggctt taatttgcaa gctgcggc
//
