LOCUS       V016083                 9876 bp    DNA     circular SYN 28-JAN-2019
DEFINITION  Exported.
ACCESSION   V016083
VERSION     V016083
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 9876)
  AUTHORS   Holwerda EK, Olson DG, Ruppertsburger NM, Stevenson D, Murphy SJL.,
            Maloney MI, Lanahan A, Amador- Noguez D, Lynd LR.
  TITLE     Evolutionary response of Clostridium thermocellum to genetic
            modifications aimed at improving ethanol production
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9876)
  AUTHORS   Olson DG, Argyros DA.
  TITLE     Direct Submission
  JOURNAL   Submitted (09-OCT-2018) Thayer School of Engineering, Dartmouth
            College, 14 Engineering Drive, Hanover, NH 03755, USA
REFERENCE   3  (bases 1 to 9876)
  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            "Unpublished"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
            (09-OCT-2018) Thayer School of Engineering, Dartmouth College, 14
            Engineering Drive, Hanover, NH 03755, USA"
FEATURES             Location/Qualifiers
     source          1..9876
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(294..1094)
                     /label="URA3"
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
     promoter        complement(1095..1315)
                     /label="URA3 promoter"
     misc_feature    1343..1846
                     /label="CEN/ARS"
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
     misc_feature    complement(1888..2922)
                     /label="3' homology region"
                     /note="3' homology region"
     misc_feature    complement(2923..3765)
                     /label="5' homology region"
                     /note="5' homology region"
     gene            complement(3884..4429)
                     /gene="hpt"
                     /label="hpt"
     CDS             complement(3884..4429)
                     /codon_start=1
                     /transl_table=11
                     /gene="hpt"
                     /product="hypoxanthine phosphoribosyltransferase"
                     /label="hpt"
                     /protein_id="QAT79155.1"
                     /translation="MENLSKDIDEILITEEELKEKIKELGRQITKDYKGKNLMLVGVLK
                     GALMFMADLSRHIDLPLSLDFMAVSSYGSSTHSSGIVKIIKDLDISIEGKDVLIVEDII
                     DSGLTLSYLRETLLGRKPKSLKICTILDKPERREASVKVDYVGFKIPDKFVVGYGLDFD
                     EKYRNLPFIGVLKPEMYS"
     CDS             complement(4455..5102)
                     /gene="cat"
                     /label="Chloramphenicol acetyltransferase"
                     /note="Chloramphenicol acetyltransferase from
                     Staphylococcus aureus. Accession#: P00485"
     regulatory      complement(5103..5630)
                     /label="gapDH promoter from Clostridium thermocellum"
                     /note="gapDH promoter from Clostridium thermocellum"
                     /regulatory_class="promoter"
     misc_feature    complement(5631..6262)
                     /label="internal homology region"
                     /note="internal homology region"
     gene            complement(6263..6841)
                     /gene="tdk"
                     /label="tdk"
     CDS             complement(6263..6841)
                     /codon_start=1
                     /transl_table=11
                     /gene="tdk"
                     /product="thymidine dikinase"
                     /label="tdk"
                     /protein_id="QAT79157.1"
                     /translation="MYGPKDHGYIEVVTGPMFSGKSEELIRRIKRAKIARQKVQVFKPA
                     IDDRYSIDKVVSHNGDNMHAIAIVKASDILAYAEEDTDVFAIDEVQFFDSEIVDIVKEI
                     ADSGKRVICAGLDMDFRGEPFGPTPELMAIAEFVDKLTAICMKCGNPATRTQRLINGKP
                     ANYDDPIIMVGAKESYEARCRKCHEVPRT"
     regulatory      complement(6842..7462)
                     /label="Clostridium thermocellum cbp promoter"
                     /note="Clostridium thermocellum cbp promoter"
                     /regulatory_class="promoter"
     CDS             complement(7572..8573)
                     /label="repB"
                     /note="RepB replication protein"
     rep_origin      complement(9265..9853)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
ORIGIN
        1 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
       61 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
      121 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
      181 agcgatctgt caattcgcgg ccgccgaaaa gtgggtaata actgatataa ttaaattgaa
      241 gctctaattt gtgagtttag tatacatgca tttacttata atacagtttt ttagttttgc
      301 tggccgcatc ttctcaaata tgcttcccag cctgcttttc tgtaacgttc accctctacc
      361 ttagcatccc ttccctttgc aaatagtcct cttccaacaa taataatgtc agatcctgta
      421 gagaccacat catccacggt tctatactgt tgacccaatg cgtctccctt gtcatctaaa
      481 cccacaccgg gtgtcataat caaccaatcg taaccttcat ctcttccacc catgtctctt
      541 tgagcaataa agccgataac aaaatctttg tcgctcttcg caatgtcaac agtaccctta
      601 gtatattctc cagtagatag ggagcccttg catgacaatt ctgctaacat caaaaggcct
      661 ctaggttcct ttgttacttc ttctgccgcc tgcttcaaac cgctaacaat acctgggccc
      721 accacaccgt gtgcattcgt aatgtctgcc cattctgcta ttctgtatac acccgcagag
      781 tactgcaatt tgactgtatt accaatgtca gcaaattttc tgtcttcgaa gagtaaaaaa
      841 ttgtacttgg cggataatgc ctttagcggc ttaactgtgc cctccatgga aaaatcagtc
      901 aagatatcca catgtgtttt tagtaaacaa attttgggac ctaatgcttc aactaactcc
      961 agtaattcct tggtggtacg aacatccaat gaagcacaca agtttgtttg cttttcgtgc
     1021 atgatattaa atagcttggc agcaacagga ctaggatgag tagcagcacg ttccttatat
     1081 gtagctttcg acatgattta tcttcgtttc ctgcaggttt ttgttctgtg cagttgggtt
     1141 aagaatactg ggcaatttca tgtttcttca acactacata tgcgtatata taccaatcta
     1201 agtctgtgct ccttccttcg ttcttccttc tgttcggaga ttaccgaatc aaaaaaattt
     1261 caaagaaacc gaaatcaaaa aaaagaataa aaaaaaaatg atgaattgaa ttgaaaagct
     1321 agcttatcga tgggtccttt tcatcacgtg ctataaaaat aattataatt taaatttttt
     1381 aatataaata tataaattaa aaatagaaag taaaaaaaga aattaaagaa aaaatagttt
     1441 ttgttttccg aagatgtaaa agactctagg gggatcgcca acaaatacta ccttttatct
     1501 tgctcttcct gctctcaggt attaatgccg aattgtttca tcttgtctgt gtagaagacc
     1561 acacacgaaa atcctgtgat tttacatttt acttatcgtt aatcgaatgt atatctattt
     1621 aatctgcttt tcttgtctaa taaatatata tgtaaagtac gctttttgtt gaaatttttt
     1681 aaacctttgt ttattttttt ttcttcattc cgtaactctt ctaccttctt tatttacttt
     1741 ctaaaatcca aatacaaaac ataaaaataa ataaacacag agtaaattcc caaattattc
     1801 catcattaaa agatacgagg cgcgtgtaag ttacaggcaa gcgatcgcgg ccgcggtacc
     1861 cggggatcct ctagagtcga cctgcagtgt catctccctt ttctgcggca tcctgaacat
     1921 ctctgaaatc actgcttaca cctgaaattc caagcacacc tgatttcttg ttaaggaaat
     1981 tgtttatatc gttaatattc attttttcct tttccatcaa ataagttata accgcagggt
     2041 caacattgcc gcttctggta cccatgcaca acccctgcag aggagtaaat cccattgagg
     2101 tgtcaacgga ttttccgcct tttaccgcac aaatacttgc tccgtttcca agatggcagg
     2161 ttatcagctt caggctctca ataggtttgc ccagcatctg agccgccctg tgggccacat
     2221 atttgtggga agttccgtgg aatccgtatt ttctcaattt atacttctca tatatctcat
     2281 aagggagggc ataaatatat gcatgccttg gcattgtctg atgaaaagct gtgtcaaaca
     2341 cagcaaccat cggcacatcg ggcagtatct gtttacaggc ttcaattccg attatatttg
     2401 acggattgtg gagcggagcc agttcaacac aatccctgat tgctttcatt acatcttcat
     2461 caattatggc agattcctta aacttctctc cgccgtgaac aatacgatga ccgacggcag
     2521 aaatttccga catgcttttt atgactccga ttttttcatc cgtaagagcc gaaattacct
     2581 cctgtatggc aagcttgtga ttgtacaggt ctttttctat aactacggtt tcccctccgg
     2641 tctttgtatg ctttaaaaag gaatgttcaa gaccaattct gtcacacaca ccttttgcaa
     2701 gcacagactc gtttgtcatg tcaatcagct gatactttag tgatgagctt ccggtattaa
     2761 taaccaaaat attcatttca aaaactcact cccgtcttgt tttttttaat tttcctattc
     2821 ctaaacttcg ataaacagat gtttttatta aacgctgcgc aacaccttct tcaatgtccg
     2881 gttttaacag aatttatgcc ttgacatatt gagcctgaac cggcggcttt aataaccctc
     2941 agatccgtac tttccggcaa aactatggtt tttacgtctg atttcgccct ttcaattatt
     3001 tgttccaaaa aactcataaa ttcttctcct ttcataatcc caaaactgtt atcataaaaa
     3061 ctgtatttgt aatacttata actatatatt atcaccaggt aataatacct actcactata
     3121 aacagctatt ttactgggtt ccaagcaact ctaattatat acaaaatgtt ttttgtatac
     3181 aacaccctcc ttatcttttt ttcggcttta gccataaata acggcaagta actccaaaat
     3241 acaggatatt tcatgctttt agaaactttt tattagtctt cttaattatt cagattttgt
     3301 ggcaattaaa ctttgcagct cctccaaata gttgtccagc tcctcttctt taagattgct
     3361 gagatatgac aatctgtaat ttttagcctt tttggccatc tctagcgcac tctccgtcat
     3421 tcccaaatct ttcaaaacac agctatagtt atagtatgcc tcaggaaaat ttggattaag
     3481 acctataagc tttcgatata tttcctcggc ctttttaatg ttaccgattt tgtaataggt
     3541 atagcctaaa ttgtcaagta tgacaggatc agagctgtta tactcatacg cctccttatt
     3601 gaaatttaaa gctttgttca tatccccgtc cagaataaga agatatccga gacttccgta
     3661 tatggttgta ttcttatagt ttgaaaacac atcctccagc atttttatgc catctttcaa
     3721 ctgtcccttt ttccaaagca ccaatgcata gttggacttt gcctggcgat aattatcagt
     3781 atccatataa cgatacctct gatcgtttta ttcaaatatg gccctccctt caaagccgta
     3841 tcacttaatt tataataaca tataccatat aaaaaaacaa attttagctg tacatttcag
     3901 gtttcaaaac gcctataaaa ggaaggttcc tgtacttttc atcaaagtcc aatccataac
     3961 ccacgacaaa cttatcaggt atcttaaatc ctacataatc gacttttaca gatgcttctc
     4021 ttctctccgg tttgtctaat attgtgcata ttttcaggct ttttggcttc cttccaagta
     4081 aagtttccct taagtaagac aaagtcaaac cgctgtcaat tatgtcttcc acaatcagaa
     4141 catctttgcc ttctatgctt atatcaagat ctttgattat ctttactatt cctgatgaat
     4201 gagttgagct tccatagctg gaaacagcca taaaatcaag tgataaaggc aaatctatgt
     4261 gtcttgacaa atcagccata aacattaaag caccttttaa aactcctacc aacatcaaat
     4321 ttttcccttt gtagtctttt gtgatttgcc tcccaagctc ttttatcttt tccttaagtt
     4381 cttcttctgt gatcaaaatt tcatcgatgt cttttgataa attttccatc aaatttcctc
     4441 cttaatcttg tttataaaag ccagtcatta ggcctatctg acaattcctg aatagagttc
     4501 ataaacaatc ctgcatgata accatcacaa acagaatgat gtacctgtaa agatagcggt
     4561 aaatatattg aattaccttt attaatgaat tttcctgctg taataatggg tagaaggtaa
     4621 ttactattat tattgatatt taagttaaac ccagtaaatg aagtccatgg aataatagaa
     4681 agagaaaaag cattttcagg tataggtgtt ttgggaaaca atttccccga accattatat
     4741 ttctctacat cagaaaggta taaatcataa aactctttga agtcattctt tacaggagtc
     4801 caaataccag agaatgtttt agatacacca tcaaaaattg tataaagtgg ctctaactta
     4861 tcccaataac ctaactctcc gtcgctattg taaccagttc taaaagctgt atttgagttt
     4921 atcacccttg tcactaagaa aataaatgca gggtaaaatt tatatccttc ttgttttatg
     4981 tttcggtata aaacactaat atcaatttct gtggttatac taaaagtcgt ttgttggttc
     5041 aaataatgat taaatatctc ttttctcttc caattgtcta aatcaatttt attaaagttc
     5101 atcattaata tcgcctccta ttgtaaatta aaatttaatt taatgcctct attagctgcc
     5161 ataaagcatg aatgcagtaa aatattcatc ctctggcaag tatctcttgc actatgattc
     5221 attttatatt attttatttt cattttcaat agtttagata aaaaataatt aattttttaa
     5281 caaagtattt ttatcttcct ttgatgtaat tctttgtttt tttgtccatt aatattatgg
     5341 acaatttttt tatttctagt caataatttt aatctttatt aaaacctggc aatgccagag
     5401 agatacagta attattgcat ataaaagaaa acagacgcgc cctgtttgtt tcctgaaaaa
     5461 ccacctcgtc ctccaaagta tccagctttt taaaatccat cctcattccc agacgaaaaa
     5521 gttttgacaa agcctctttt cttgtccata gaattgtact ctgcacacaa tattcatcag
     5581 tatttccaaa actctttaga atcttttcct ctctttcgga aaagaaatac caatggcggc
     5641 atccacctga agttctccgt caattgcaag gtggggagct ttttcctttg caagctgtgt
     5701 tgccttgatt accttttcgg tcagctcact tttggcactg ccgtaagaag aataagaaag
     5761 cattgccacc tgaggttttg ctccaaccag catctcaaaa gatttggatg cagatattgc
     5821 aatttcagaa agctggtctg catccggatt ttccaccaag ccgcaatcgg catatacaaa
     5881 ggttccgtta tgaccatatt cacagttggg tacaaccata acaaaaaagg atgatacgag
     5941 ttttgtcccc ggggccgtct ttaatatctg caaagccggt ctcaaagtat ttgcagtgga
     6001 attgacagca cccgccacca taccatccgc ttcacctttt tttaccatca taactccata
     6061 ataaagaggg tctttgatcg tttcccttgc ggcttctata gtcataccct tcgattttct
     6121 aagctcatac agtgtatttg cataatcctc caatttttcg gaatttaagg aatcctctat
     6181 catcactcct tcaagatcaa tatcccccgc cagactctta atctcctttt cattgcctat
     6241 cagtacaacc tttgcaattc ccttaagtcc gcgggacttc atggcacttt ctacaccttg
     6301 cttcataaga ctcctttgct ccaaccatta taatgggatc gtcgtaattg gcaggcttcc
     6361 catttataag cctttgcgtg cgagtagcag gattgccaca cttcatgcat atagcagtaa
     6421 gcttatcgac aaattcagct atggccatca attctggagt cggaccaaat ggttcacctc
     6481 taaagtccat gtcaagccct gcacatatga ctcttttccc gctatcggca atctctttta
     6541 cgatgtcgac tatttcagaa tcaaaaaatt gaacttcatc tatagcaaat acatccgtat
     6601 cttcttcagc ataagccaat atgtcagaag cctttactat ggcaatggcg tgcatgttgt
     6661 cgccgttatg agatacgact ttgtctatgg aatacctatc gtctatagcc ggtttgaaaa
     6721 cctgaacttt ttgcctggca atcttagctc tctttatcct tcttataagt tcctcacttt
     6781 ttccactgaa catgggacca gttacaactt ctatgtaccc gtggtcttta ggcccataca
     6841 tctttcgtcc tccttaaaat tttcgtttat ttgtttttta tttagtgtgc ctaaaagcac
     6901 catcttatac attcgtacct tatatctctt atcttgtatc tcttacctca tatctcttat
     6961 tattaactac cgaagtatat tttcataacg gaatcttatg acttattttt ctgactgccg
     7021 ttttaccata cggttaattt atacggaaac agctaatcta accttgctga aggcatgttt
     7081 accaaaatat tatatctact ttaccatatt aattcaagtg gtcattttga tattttacgc
     7141 caaaattttt agcgttcctt tacttgataa tttatccaaa aaccttcatt cgtcaaaaca
     7201 attatgatat aattaattta tgatatgatt aattaatatt tacggataac taatcatttc
     7261 aatgcaatag atcaacatgc aaatcaatat ccaccttact ccgaacattt tgcaccaacg
     7321 aatgcccagc actatatccc ggcaagcaat ataacgtcta taatttttta tgataacgat
     7381 aaaattaaat tttatgaaag gggtaaaagc atgagaaaaa tagctgtatt gttaattact
     7441 ttgacgttct tagtcacgac tcttgatatg cctcctaaat ttttatctaa agtgaattta
     7501 ggaggcttac ttgtctgctt tcttcattag aatcaatcct tttttaaaag tcaatcccgt
     7561 ttgttgaact actctttaat aaaataattt ttccgttccc aattccacat tgcaataata
     7621 gaaaatccat cttcatcggc tttttcgtca tcatctgtat gaatcaaatc gccttcttct
     7681 gtgtcatcaa ggtttaattt tttatgtatt tcttttaaca aaccaccata ggagattaac
     7741 cttttacggt gtaaaccttc ctccaaatca gacaaacgtt tcaaattctt ttcttcatca
     7801 tcggtcataa aatccgtatc ctttacagga tattttgcag tttcgtcaat tgccgattgt
     7861 atatccgatt tatatttatt tttcggtcga atcatttgaa cttttacatt tggatcatag
     7921 tctaatttca ttgccttttt ccaaaattga atccattgtt tttgattcac gtagttttct
     7981 gtattcttaa aataagttgg ttccacacat accaatacat gcatgtgctg attataagaa
     8041 ttatctttat tatttattgt cacttccgtt gcacgcataa aaccaacaag atttttatta
     8101 atttttttat attgcatcat tcggcgaaat ccttgagcca tatctgacaa actcttattt
     8161 aattcttcgc catcataaac atttttaact gttaatgtga gaaacaacca acgaactgtt
     8221 ggcttttgtt taataacttc agcaacaacc ttttgtgact gaatgccatg tttcattgct
     8281 ctcctccagt tgcacattgg acaaagcctg gatttacaaa accacactcg atacaacttt
     8341 ctttcgcctg tttcacgatt ttgtttatac tctaatattt cagcacaatc ttttactctt
     8401 tcagcctttt taaattcaag aatatgcaga agttcaaagt aatcaacatt agcgattttc
     8461 ttttctctcc atggtctcac ttttccactt tttgtcttgt ccactaaaac ccttgatttt
     8521 tcatctgaat aaatgctact attaggacac ataatattaa aagaaacccc catctattta
     8581 gttatttgtt tggtcactta taactttaac agatggggtt tttctgtgca accaatttta
     8641 agggttttca atactttaaa acacatacat accaacactt caacgcacct ttcagcaact
     8701 aaaataaaaa tgacgttatt tctatatgta tcaagaatag aaagaactcg tttttcgcta
     8761 cgctcaaaac gcaaaaaaag cactcattcg agtgcttttt cttatcgctc caaatcatgc
     8821 gattttttcc tctttgcttt tctttgctca cgaagttctc gatcacgctg caaaacatct
     8881 tgaagcgaaa aagtattctt cttttcttcc gatcgctcat gctgacgcac gaaaagccct
     8941 ctaggcgcat aggaacaact cctaaatgca tgtgaggggt tttctcgtcc atgtgaacag
     9001 tcgcatacgc aatattttgt ttcccatact gcattaatga atcggccaac gcgcggggag
     9061 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     9121 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     9181 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     9241 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     9301 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     9361 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     9421 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     9481 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     9541 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     9601 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     9661 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     9721 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     9781 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     9841 aaaaggatct caagaagatc ctttgatctt ttctac
//
