LOCUS       V015730                 8469 bp    DNA     circular SYN 05-OCT-2020
DEFINITION  Exported.
ACCESSION   V015730
VERSION     V015730
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8469)
  AUTHORS   Holwerda EK, Zhou J, Hon S, Stevenson D, Amador-Noguez D, Lynd LR,
            van Dijken JP.
  TITLE     Metabolic fluxes of nitrogen and pyrophosphate in chemostat cultures
            of Clostridium thermocellum and Thermoanaerobacterium
            saccharolyticum
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8469)
  AUTHORS   Hon S.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-APR-2020) Thayer School of Engineering, Dartmouth
            College, 14 Engineering Drive, Hanover, NH 03755, USA
REFERENCE   3  (bases 1 to 8469)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing
            ##Assembly-Data-END##
            SGRef: number: 1; type: "Journal Article"; journalName:
            "Unpublished"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (28-APR-2020) Thayer School of Engineering, Dartmouth College, 14
            Engineering Drive, Hanover, NH 03755, USA"
FEATURES             Location/Qualifiers
     source          1..8469
                     /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."
     gene            complement(840..1418)
                     /gene="tdk"
                     /label="tdk"
     CDS             complement(840..1418)
                     /codon_start=1
                     /transl_table=11
                     /gene="tdk"
                     /product="thymidine kinase"
                     /label="tdk"
                     /protein_id="QOE54900.1"
                     /translation="MYGPKDHGYIEVVTGPMFSGKSEELIRRIKRAKIARQKVQVFKPA
                     IDDRYSIDKVVSHNGDNMHAIAIVKASDILAYAEEDTDVFAIDEVQFFDSEIVDIVKEI
                     ADSGKRVICAGLDMDFRGEPFGPTPELMAIAEFVDKLTAICMKCGNPATRTQRLINGKP
                     ANYDDPIIMVGAKESYEARCRKCHEVPRT"
     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..5244
                     /label="upstream homologous recombination flank"
                     /note="upstream homologous recombination flank"
     misc_feature    5245..5736
                     /label="downstream homologous recombination flank"
                     /note="downstream homologous recombination flank"
     regulatory      5740..6314
                     /label="C. thermocellum gapdh promoter"
                     /note="C. thermocellum gapdh promoter"
                     /regulatory_class="promoter"
     CDS             6315..6962
                     /gene="cat"
                     /label="Chloramphenicol acetyltransferase"
                     /note="Chloramphenicol acetyltransferase from
                     Staphylococcus aureus. Accession#: P00485"
     gene            6983..7528
                     /gene="hpt"
                     /label="hpt"
     CDS             6983..7528
                     /codon_start=1
                     /transl_table=11
                     /gene="hpt"
                     /product="hypoxanthine phosphoribosyltransferase"
                     /label="hpt"
                     /protein_id="QOE54904.1"
                     /translation="MENLSKDIDEILITEEELKEKIKELGRQITKDYKGKNLMLVGVLK
                     GALMFMADLSRHIDLPLSLDFMAVSSYGSSTHSSGIVKIIKDLDISIEGKDVLIVEDII
                     DSGLTLSYLRETLLGRKPKSLKICTILDKPERREASVKVDYVGFKIPDKFVVGYGLDFD
                     EKYRNLPFIGVLKPEMYS"
     misc_feature    7713..8226
                     /note="gene target internal homologous recombination
                     region"
     terminator      8259..8447
                     /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 tgacatactt
     4741 ctgaaaatgg ctttgaaggc aaatatcccg ggattatcat tcttcttatt gtaatcatta
     4801 tccttctgct cattacagat gtttatatat tagtgtcaga caaagaaaag ctaaaaagtt
     4861 ttaagctgtt taagcgacta aaatgacggg aaatagaaaa aaataatcgc agtattataa
     4921 ttttttacag cctcttatga tataatagcg aaggaaaaat ataatagcga aggaaaaaaa
     4981 tttaaataaa ctgcggacgt tatttttaaa tttatcataa ttttaaagcc ataataataa
     5041 aaaacagaag ggaggtttat tttaaaaaag cagttgtaag gttgcttccg gtatttagtc
     5101 aggtgcttaa tcaaaaagca aattttattt tcgtttcagc ttttcatgtt atgttcatca
     5161 aatgaaaagc cgaaataaga tttaagtttt atgtaattgc tgttttgatg taaaagaatc
     5221 taagttcacc aaggaggtct gtaaaaagcc ttcctttcac ggaaggcttt ttgcgttttt
     5281 tgcttctaag actaaaacaa aacgttcaat atttcttaca ctaccggcgt ataatttcga
     5341 ctcaaactgt tcgaatatat ccttatattt gcataaaata tgaaacagtc acatcagcct
     5401 aaactgcgtg ccctcagtgt ttcactattt ttacttcatt ttttcaatct gtttttcaat
     5461 caaaaacact atagatattt tttcaattta gtattataat gttctaatgt ttgataagaa
     5521 tatcacattc aaactaactt gaaaatgttt ttattgagga gtttttattc atgaacaaaa
     5581 gtcaaacaaa aataaatgat tttgtctatg gatttaaaaa aggaacaccg attctattag
     5641 gatacatacc ggtagctttt actttcggcc ttatggctgt cggcggcgga cttcatccgt
     5701 taatggcagt gtttatttcc ctgaccaacc ttaccaatcg tgggaatagg catggaaaag
     5761 atttttttgc ccgaggatgc attgataaag tatttctttt ccgaaagaga ggaaaagatt
     5821 ctaaagagtt ttggaaatac tgatgaatat tgtgtgcaga gtacaattct atggacaaga
     5881 aaagaggctt tgtcaaaact ttttcgtctg ggaatgagga tggattttaa aaagctggat
     5941 actttggagg gcgaggtggt ttttcaggaa acaaacaggg cgcgtctgtt ttcttttata
     6001 tgcaataatt actgtatctc tctggcattg ccaggtttta ataaagatta aaattattga
     6061 ctagaaataa aaaaattgtc cataatatta atggacaaaa aaacaaagaa ttacatcaaa
     6121 ggaagataaa aatactttgt taaaaaatta attatttttt atctaaacta ttgaaaatga
     6181 aaataaaata atataaaatg aatcatagtg caagagatac ttgccagagg atgaatattt
     6241 tactgcattc atgctttatg gcagctaata gaggcattaa attaaatttt aatttacaat
     6301 aggaggcgat attaatgaac tttaataaaa ttgatttaga caattggaag agaaaagaga
     6361 tatttaatca ttatttgaac caacaaacga cttttagtat aaccacagaa attgatatta
     6421 gtgttttata ccgaaacata aaacaagaag gatataaatt ttaccctgca tttattttct
     6481 tagtgacaag ggtgataaac tcaaatacag cttttagaac tggttacaat agcgacggag
     6541 agttaggtta ttgggataag ttagagccac tttatacaat ttttgatggt gtatctaaaa
     6601 cattctctgg tatttggact cctgtaaaga atgacttcaa agagttttat gatttatacc
     6661 tttctgatgt agagaaatat aatggttcgg ggaaattgtt tcccaaaaca cctatacctg
     6721 aaaatgcttt ttctctttct attattccat ggacttcatt tactgggttt aacttaaata
     6781 tcaataataa tagtaattac cttctaccca ttattacagc aggaaaattc attaataaag
     6841 gtaattcaat atatttaccg ctatctttac aggtacatca ttctgtttgt gatggttatc
     6901 atgcaggatt gtttatgaac tctattcagg aattgtcaga taggcctaat gactggcttt
     6961 tataataaag gaggtcgacg tcatggaaaa tttatcaaaa gacatcgatg aaattttgat
     7021 cacagaagaa gaacttaagg aaaagataaa agagcttggg aggcaaatca caaaagacta
     7081 caaagggaaa aatttgatgt tggtaggagt tttaaaaggt gctttaatgt ttatggctga
     7141 tttgtcaaga cacatagatt tgcctttatc acttgatttt atggctgttt ccagctatgg
     7201 aagctcaact cattcatcag gaatagtaaa gataatcaaa gatcttgata taagcataga
     7261 aggcaaagat gttctgattg tggaagacat aattgacagc ggtttgactt tgtcttactt
     7321 aagggaaact ttacttggaa ggaagccaaa aagcctgaaa atatgcacaa tattagacaa
     7381 accggagaga agagaagcat ctgtaaaagt cgattatgta ggatttaaga tacctgataa
     7441 gtttgtcgtg ggttatggat tggactttga tgaaaagtac aggaaccttc cttttatagg
     7501 cgttttgaaa cctgaaatgt acagctaagc ccgggcctcg agaaaacaaa aggctcagtc
     7561 ggaagactgg gccttttgtt ttggtaccga attcggcgcg cctcagcgtt taaacccgct
     7621 gatcctagag ggccgcatca tgtaattagt tatgtcacgc ttacattcac gccctcccca
     7681 tgctttaata catacctggc ccagtagttc aggacgatcc gagaaatccc gcagttatag
     7741 ccgacaacgt cggagacaac gtaggtgatg ttgccggaat gggcgcagac ctcttcgact
     7801 ccaacgttgc ctcaatggcc gctgcactgg ttatggccac tgctctcgga ggacctgaca
     7861 gcaaaaacgt agcaatggtt ttcgcctatg cggcaatagg tcttttgtct tcggttattg
     7921 gagtggctac ggcacgcatc ggcaaaaacg gtgacccggg aaaagctctc aacagttcca
     7981 cttatgtgac caccgccatt tttgccgtac ttacggcttt gtcaacctac atctttaact
     8041 tcgaatggcg tatttggggc gcaactgcca taggtctgat agtcggaacc attataggtc
     8101 ttacctctga ctatttcaca aacgacgcaa aacctccggt tcgagccgtt gcaaatgctt
     8161 caaagtcagg acctgctttt accatacttt caggtttttc ctacggattg ctaagtgccc
     8221 ttccttctgc taatagtagt gaaaaaatca gggataatgc cgctctaacc gaaaaggaag
     8281 gagttagaca acctgaagtc taggtcccta tttatttttt tatagttatg ttagtattaa
     8341 gaacgttatt tatatttcaa atttttcttt tttttctgta cagacgcgtg tacgcatgta
     8401 acattatact gaaaaccttg cttgagaagg ttttgggacg ctcgaaggct ttaatttgca
     8461 agctgcggc
//
