LOCUS       V008010                10709 bp    DNA     circular SYN 18-DEC-2018
DEFINITION  Exported.
ACCESSION   V008010
VERSION     V008010
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 10709)
  AUTHORS   Olson DG, Tripathi SA, Giannone RJ, Lo J, Caiazza NC, Hogsett DA,
            Hettich RL, Guss AM, Dubrovsky G, Lynd LR.
  TITLE     Deletion of the Cel48S cellulase from Clostridium thermocellum
  JOURNAL   Proc. Natl. Acad. Sci. U.S.A. 107 (41), 17727-17732 (2010)
   PUBMED   20837514
REFERENCE   2  (bases 1 to 10709)
  AUTHORS   Olson DG.
  TITLE     Direct Submission
  JOURNAL   Submitted (18-AUG-2010) Thayer School of Engineering, Dartmouth
            College, 8000 Cummings, Hanover, NH 03755, USA
REFERENCE   3  (bases 1 to 10709)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 10709)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proc. Natl.
            Acad. Sci. U.S.A."; date: "2010"; volume: "107"; issue: "41"; pages:
            "17727-17732"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (18-AUG-2010) Thayer School of Engineering, Dartmouth College, 8000
            Cummings, Hanover, NH 03755, USA"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..10709
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             126..926
                     /label="URA3"
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
     rep_origin      1244..1832
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             2524..3525
                     /label="repB"
                     /note="RepB replication protein"
     regulatory      3529..4149
                     /note="cbp; derived from Clostridium thermocellum strain
                     DSM 1313"
                     /regulatory_class="promoter"
     CDS             4150..5094
                     /codon_start=1
                     /product="pyrF"
                     /label="pyrF"
                     /note="negative-selectable marker used with 5FOA"
                     /protein_id="ADW95856.1"
                     /translation="MFIDTLIEKIREKDNPSVVGLDPKIEYVPSFIKEDMYKKYGKNLK
                     AVAEAILLFNKYIIDAVYDIVPAVKPQLAYYEMYGLEGMRVFYETCKYAKEKGLLVIAD
                     GKRNDIGSTAQCYSAAYLGKTDIDEGISEAVFDVDALTVNPYLGVDGIKPFIDDCVKYN
                     KGIFVLVKTSNKSSGEIQDILTQEGRSIYEIVAEYVESWGENKKGKYGYSCVGAVVGAT
                     YPNLAKILRKILKNSYILVPGYGAQGGTARDVAHCFNYDGLGAIVNASRSIMCAYKSEQ
                     WKNVYSEEKFYEASRAEAIRMRDDINSALRDRK"
     terminator      5220..5301
                     /label="rrnB T1 terminator"
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      5393..5420
                     /label="rrnB T2 terminator"
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     promoter        5501..5605
                     /label="AmpR promoter"
     CDS             5606..6463
                     /label="AmpR"
                     /note="beta-lactamase"
     protein_bind    6625..6641
                     /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)."
     primer_bind     6649..6665
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     misc_feature    complement(6710..7767)
                     /note="cel48S 3' flanking region; derived from Clostridium
                     thermocellum strain DSM 1313"
     regulatory      7841..8365
                     /note="gapDH; derived from Clostridium thermocellum strain
                     DSM 1313"
                     /regulatory_class="promoter"
     CDS             8366..9013
                     /gene="cat"
                     /label="Chloramphenicol acetyltransferase"
                     /note="Chloramphenicol acetyltransferase from
                     Staphylococcus aureus. Accession#: P00485"
     misc_feature    complement(9097..10064)
                     /note="cel48S 5' flanking region; derived from Clostridium
                     thermocellum strain DSM 1313"
     misc_feature    complement(10083..10586)
                     /label="CEN/ARS"
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
ORIGIN
        1 gaaggaagga gcacagactt agattggtat atatacgcat atgtagtgtt gaagaaacat
       61 gaaattgccc agtattctta acccaactgc acagaacaaa aacctgcagg aaacgaagat
      121 aaatcatgtc gaaagctaca tataaggaac gtgctgctac tcatcctagt cctgttgctg
      181 ccaagctatt taatatcatg cacgaaaagc aaacaaactt gtgtgcttca ttggatgttc
      241 gtaccaccaa ggaattactg gagttagttg aagcattagg tcccaaaatt tgtttactaa
      301 aaacacatgt ggatatcttg actgattttt ccatggaggg cacagttaag ccgctaaagg
      361 cattatccgc caagtacaat tttttactct tcgaagacag aaaatttgct gacattggta
      421 atacagtcaa attgcagtac tctgcgggtg tatacagaat agcagaatgg gcagacatta
      481 cgaatgcaca cggtgtggtg ggcccaggta ttgttagcgg tttgaagcag gcggcagaag
      541 aagtaacaaa ggaacctaga ggccttttga tgttagcaga attgtcatgc aagggctccc
      601 tatctactgg agaatatact aagggtactg ttgacattgc gaagagcgac aaagattttg
      661 ttatcggctt tattgctcaa agagacatgg gtggaagaga tgaaggttac gattggttga
      721 ttatgacacc cggtgtgggt ttagatgaca agggagacgc attgggtcaa cagtatagaa
      781 ccgtggatga tgtggtctct acaggatctg acattattat tgttggaaga ggactatttg
      841 caaagggaag ggatgctaag gtagagggtg aacgttacag aaaagcaggc tgggaagcat
      901 atttgagaag atgcggccag caaaactaaa aaactgtatt ataagtaaat gcatgtatac
      961 taaactcaca aattagagct tcaatttaat tatatcagtt attacccact tttcggcggc
     1021 cgcgaattga cagatcgctg agataggtgc ctcactggat taagcattgg taactgtcag
     1081 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     1141 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     1201 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     1261 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     1321 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     1381 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     1441 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     1501 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     1561 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     1621 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     1681 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     1741 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     1801 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     1861 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     1921 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     1981 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc
     2041 ccgcgcgttg gccgattcat taatgcagta tgggaaacaa aatattgcgt atgcgactgt
     2101 tcacatggac gagaaaaccc ctcacatgca tttaggagtt gttcctatgc gcctagaggg
     2161 cttttcgtgc gtcagcatga gcgatcggaa gaaaagaaga atactttttc gcttcaagat
     2221 gttttgcagc gtgatcgaga acttcgtgag caaagaaaag caaagaggaa aaaatcgcat
     2281 gatttggagc gataagaaaa agcactcgaa tgagtgcttt ttttgcgttt tgagcgtagc
     2341 gaaaaacgag ttctttctat tcttgataca tatagaaata acgtcatttt tattttagtt
     2401 gctgaaaggt gcgttgaagt gttggtatgt atgtgtttta aagtattgaa aacccttaaa
     2461 attggttgca cagaaaaacc ccatctgtta aagttataag tgaccaaaca aataactaaa
     2521 tagatggggg tttcttttaa tattatgtgt cctaatagta gcatttattc agatgaaaaa
     2581 tcaagggttt tagtggacaa gacaaaaagt ggaaaagtga gaccatggag agaaaagaaa
     2641 atcgctaatg ttgattactt tgaacttctg catattcttg aatttaaaaa ggctgaaaga
     2701 gtaaaagatt gtgctgaaat attagagtat aaacaaaatc gtgaaacagg cgaaagaaag
     2761 ttgtatcgag tgtggttttg taaatccagg ctttgtccaa tgtgcaactg gaggagagca
     2821 atgaaacatg gcattcagtc acaaaaggtt gttgctgaag ttattaaaca aaagccaaca
     2881 gttcgttggt tgtttctcac attaacagtt aaaaatgttt atgatggcga agaattaaat
     2941 aagagtttgt cagatatggc tcaaggattt cgccgaatga tgcaatataa aaaaattaat
     3001 aaaaatcttg ttggttttat gcgtgcaacg gaagtgacaa taaataataa agataattct
     3061 tataatcagc acatgcatgt attggtatgt gtggaaccaa cttattttaa gaatacagaa
     3121 aactacgtga atcaaaaaca atggattcaa ttttggaaaa aggcaatgaa attagactat
     3181 gatccaaatg taaaagttca aatgattcga ccgaaaaata aatataaatc ggatatacaa
     3241 tcggcaattg acgaaactgc aaaatatcct gtaaaggata cggattttat gaccgatgat
     3301 gaagaaaaga atttgaaacg tttgtctgat ttggaggaag gtttacaccg taaaaggtta
     3361 atctcctatg gtggtttgtt aaaagaaata cataaaaaat taaaccttga tgacacagaa
     3421 gaaggcgatt tgattcatac agatgatgac gaaaaagccg atgaagatgg attttctatt
     3481 attgcaatgt ggaattggga acggaaaaat tattttatta aagagtagga gtcgtgacta
     3541 agaacgtcaa agtaattaac aatacagcta tttttctcat gcttttaccc ctttcataaa
     3601 atttaatttt atcgttatca taaaaaatta tagacgttat attgcttgcc gggatatagt
     3661 gctgggcatt cgttggtgca aaatgttcgg agtaaggtgg atattgattt gcatgttgat
     3721 ctattgcatt gaaatgatta gttatccgta aatattaatt aatcatatca taaattaatt
     3781 atatcataat tgttttgacg aatgaaggtt tttggataaa ttatcaagta aaggaacgct
     3841 aaaaattttg gcgtaaaata tcaaaatgac cacttgaatt aatatggtaa agtagatata
     3901 atattttggt aaacatgcct tcagcaaggt tagattagct gtttccgtat aaattaaccg
     3961 tatggtaaaa cggcagtcag aaaaataagt cataagattc cgttatgaaa atatacttcg
     4021 gtagttaata ataagagata tgaggtaaga gatacaagat aagagatata aggtacgaat
     4081 gtataagatg gtgcttttag gcacactaaa taaaaaacaa ataaacgaaa attttaagga
     4141 ggacgaaaga tgtttattga tacattaatt gaaaagatta gagaaaagga taatccttcc
     4201 gttgtaggat tagaccctaa aattgaatat gttccgtctt ttataaagga agacatgtat
     4261 aaaaaatacg ggaaaaattt aaaagctgtg gcagaggcga ttctcctctt caataaatat
     4321 attattgatg cggtttacga tattgttcct gcagtaaaac cgcagcttgc atattatgaa
     4381 atgtacggcc ttgaaggcat gagggtgttt tatgaaactt gcaaatatgc aaaggaaaaa
     4441 ggacttttgg ttattgcaga cggaaaaaga aacgacatag gttccaccgc ccagtgttat
     4501 tctgccgcat atcttggaaa aacggacatt gatgaaggta taagcgaggc ggtttttgat
     4561 gtggatgccc tgacagtcaa cccgtatctt ggtgtggacg gtattaagcc ttttatagat
     4621 gactgtgtca agtacaacaa gggcatattt gttctggtca agacatcaaa caagtcatcc
     4681 ggagaaattc aggacatact cacccaggaa ggaagaagca tttatgagat tgttgcggag
     4741 tatgttgaat catggggtga aaacaaaaaa ggaaaatatg gatacagttg tgtgggagca
     4801 gtggttggag caacttatcc caatctggcc aaaattttaa gaaagattct gaaaaattcc
     4861 tatatactgg ttccgggcta tggagctcag ggaggaacag ccagagatgt agcccattgc
     4921 tttaattatg acgggctcgg agcaattgtc aatgcatcaa gaagcataat gtgtgcctac
     4981 aaatctgaac aatggaagaa tgtttacagc gaagaaaagt tttatgaggc atcaagagcc
     5041 gaggcaataa gaatgaggga cgatattaac agtgcgttgc gagacaggaa gtaataaagc
     5101 aaggtatgac aaaaagaagt tatccgggtt tttcggataa cttctttttt aaaatagcgg
     5161 cggttaaaac tctattcaat cctgcgttct tgaattcgcg aactcagaag tgaaacgata
     5221 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     5281 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
     5341 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc
     5401 atcctgacgg atggcctttt ttattgtaaa ttccggtaac ccttgttcta gagtcgacct
     5461 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     5521 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     5581 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     5641 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     5701 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     5761 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     5821 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     5881 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     5941 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     6001 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     6061 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     6121 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     6181 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     6241 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     6301 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     6361 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     6421 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     6481 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     6541 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     6601 agcgtcaaga tctgttgtgt ggaattgtga gcggataaca atttcacaca ggaaacagct
     6661 atgaccatga ttacgccaag cttgcatgcc ctgcaggtcg actctagagc ggtattttct
     6721 ctcctttggg tctttgattt tttcaatata cggcgttctg tacacattcg aatttgttac
     6781 ggaaaaacct ctttttataa gctctttaaa gaggttgttt tcaattatca tcttcttcag
     6841 tgtaattgtg ggttgtccgt gcaaatgcct ggacaggacc tttgacgcat tttgaccggt
     6901 aaatatggtt gtctgccccg ttgcactctg gggcaggccg ggcacgtcaa gacaggcatc
     6961 cgtaggaatt gcaccggctt tattaagtat ataatttatg tttgggagct ccggattggc
     7021 cagaggattg accgaagcat cctcctttcc aataccaaaa ccatctataa agatgaatat
     7081 cattttcacg gttggacacc accctgtaaa aaagtaaaaa gaagcgccct ttacgcttcc
     7141 ctgccattta accgacatcc catattttaa attcttgctc tcgttgactt ctcaagggta
     7201 tttcgaactt cattcaactc agcttctctc tttgccagct ccaattgata gtaggtattt
     7261 ttgttgctta ttacggcatt ttcattggcg agcctccggt tttcctcttt cagcctttct
     7321 atctcttcct ttgcactttt caactccctg ctgagacttg caatctcctc ctgagctttg
     7381 aaatactcat cggcaacatt aacggcggta agcaccgatg ccattgaggt gctaagctta
     7441 ttattcattc tcatgatttc agtcatcttt ttgtctatat acagcgctac cttctgaata
     7501 tactcttcgg attcgcaacc aaccaacgta taatcttttc ccgctatcct gacttccacc
     7561 ttgtttttct ccgccatacc atcagctcct ttgcacattg cagaccatgc gactcttttt
     7621 tcaaaaaaga acacattatt atttattata caatatttcg gcacaaaaat cattggtttt
     7681 tagcgaatat aatagaaatt atgcgaaaag ccatggtatg gctttagaaa tgtaaaaagg
     7741 accggcacaa gccggtcctt tcaaatcgta gttcaacaaa cgggattgac ttttaaaaaa
     7801 ggattgattc taatgaagaa agcagacaag taagcctcct gtatttcttt tccgaaagag
     7861 aggaaaagat tctaaagagt tttggaaata ctgatgaata ttgtgtgcag agtacaattc
     7921 tatggacaag aaaagaggct ttgtcaaaac tttttcgtct gggaatgagg atggatttta
     7981 aaaagctgga tactttggag ggcgaggtgg tttttcagga aacaaacagg gcgcgtctgt
     8041 tttcttttat atgcaataat tactgtatct ctctggcatt gccaggtttt aataaagatt
     8101 aaaattattg actagaaata aaaaaattgt ccataatatt aatggacaaa aaaacaaaga
     8161 attacatcaa aggaagataa aaatactttg ttaaaaaatt aattattttt tatctaaact
     8221 attgaaaatg aaaataaaat aatataaaat gaatcatagt gcaagagata cttgccagag
     8281 gatgaatatt ttactgcatt catgctttat ggcagctaat agaggcatta aattaaattt
     8341 taatttacaa taggaggcga tattaatgaa ctttaataaa attgatttag acaattggaa
     8401 gagaaaagag atatttaatc attatttgaa ccaacaaacg acttttagta taaccacaga
     8461 aattgatatt agtgttttat accgaaacat aaaacaagaa ggatataaat tttaccctgc
     8521 atttattttc ttagtgacaa gggtgataaa ctcaaataca gcttttagaa ctggttacaa
     8581 tagcgacgga gagttaggtt attgggataa gttagagcca ctttatacaa tttttgatgg
     8641 tgtatctaaa acattctctg gtatttggac tcctgtaaag aatgacttca aagagtttta
     8701 tgatttatac ctttctgatg tagagaaata taatggttcg gggaaattgt ttcccaaaac
     8761 acctatacct gaaaatgctt tttctctttc tattattcca tggacttcat ttactgggtt
     8821 taacttaaat atcaataata atagtaatta ccttctaccc attattacag caggaaaatt
     8881 cattaataaa ggtaattcaa tatatttacc gctatcttta caggtacatc attctgtttg
     8941 tgatggttat catgcaggat tgtttatgaa ctctattcag gaattgtcag ataggcctaa
     9001 tgactggctt ttataatatg agataatgcc gactgtactt tttacagtcg gttttctaat
     9061 gtcactaggg ctcgcctttg ggaagtttga agggctatga aataatgcaa cctacccgca
     9121 ataaattcat tccccctttt tgcggataaa ttttacgcat tattttcatc ataaaactaa
     9181 gctggaccca aaaaatactt gtgttaacat aataaatttg tgttaacaaa cttttttatc
     9241 aaatcagccg cgcatttgac ataaatttac atttttttgt cggaacgacg tataacaaaa
     9301 gaaactgcca cgaagacaat actgaaaggt ttaaaagaca attgcttttt tgaacagaga
     9361 ctattgcaaa tactgaaatt tgtagatgcc agattattac tacaataaat acatcttatt
     9421 atgggaattt aaacagaccg gcgaagtctt acgtcaaatg cagctgaatc acgcaaaaca
     9481 acttaacata ttcagttttc aacgacccta aaaggccaaa attaataact taaattaatc
     9541 aataaattcc atatatatat cttcgaaaaa aatatacttt ttgagggggc aacaaagaaa
     9601 tctccaattt ttaaatttca ctatggaaac atttgaagtt cttgtgcttt tttcgaaaga
     9661 cagcagatat gcaaccccat tatacgatat taatactacc cccatttcaa aaataactga
     9721 gtaaattatt gctattttgt tttgtatttt tattataata cacatcaagt ttgtatgcaa
     9781 gtcataaaat ttatatttaa ccaagtctga ctttaatagt actacaaaca tgaaataacg
     9841 gtgaacccgg cctgccaaaa ataaaaaagc ctacggcgca gttaaaagcc atagactaaa
     9901 aatatcttgc caaagaatat taaaatatta aaacttcatt tttcctcgtt ttgctcagcg
     9961 gggcattcgc tcccgcagct ttcttccttg ttctctccgg tagtttcctt gctgcactca
    10021 acattatttt ccacattact ttccgcatta cattcgggat tacacccggg taccgcggcc
    10081 gcgatcgctt gcctgtaact tacacgcgcc tcgtatcttt taatgatgga ataatttggg
    10141 aatttactct gtgtttattt atttttatgt tttgtatttg gattttagaa agtaaataaa
    10201 gaaggtagaa gagttacgga atgaagaaaa aaaaataaac aaaggtttaa aaaatttcaa
    10261 caaaaagcgt actttacata tatatttatt agacaagaaa agcagattaa atagatatac
    10321 attcgattaa cgataagtaa aatgtaaaat cacaggattt tcgtgtgtgg tcttctacac
    10381 agacaagatg aaacaattcg gcattaatac ctgagagcag gaagagcaag ataaaaggta
    10441 gtatttgttg gcgatccccc tagagtcttt tacatcttcg gaaaacaaaa actatttttt
    10501 ctttaatttc tttttttact ttctattttt aatttatata tttatattaa aaaatttaaa
    10561 ttataattat ttttatagca cgtgatgaaa aggacccatc gataagctag cttttcaatt
    10621 caattcatca tttttttttt attctttttt ttgatttcgg tttctttgaa atttttttga
    10681 ttcggtaatc tccgaacaga aggaagaac
//
