LOCUS       40924_48473       10036 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pZJ16, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10036)
  AUTHORS   Zhou J, Olson DG, Lanahan AA, Tian L, Murphy SJ, Lo J, Lynd LR.
  TITLE     Physiological roles of pyruvate ferredoxin oxidoreductase and 
            pyruvate formate-lyase in Thermoanaerobacterium saccharolyticum 
            JW/SL-YS485
  JOURNAL   Biotechnol Biofuels 8, 138 (2015)
  PUBMED    26379770
REFERENCE   2  (bases 1 to 10036)
  AUTHORS   Zhou J.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-OCT-2014) Thayer School of Engineering, Dartmouth 
            College, 14 Engineering Drive, Hanover, NH 03755, USA
REFERENCE   3  (bases 1 to 10036)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 10036)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Biotechnol 
            Biofuels 8, 138 (2015)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (27-OCT-2014) Thayer School of Engineering, Dartmouth College, 14 
            Engineering Drive, Hanover, NH 03755, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..10036
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            complement(347..456)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     terminator      complement(877..904)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     terminator      complement(996..1082)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     misc_feature    1144..2123
                     /label=upstream of Tsac_1064
                     /note="upstream of Tsac_1064"
     CDS             2273..3331
                     /codon_start=1
                     /gene="pta"
                     /product="phosphate acetyltransferase"
                     /label=pta
                     /note="negative selective marker in Thermoanaerobacter 
                     saccharolyticum"
                     /protein_id="AJP77036.1"
                     /translation="MYTIYFFLVRGMYKNKYFKGRDDLMSIIQNIIEKAKSDKKKIVLP
                     EGAEPRTLKAAEIVLKEGIADLVLLGNEDEIRNAAKDLDISKAEIIDPVKSEMFDRYAN
                     DFYELRKNKGITLEKARETIKDNIYFGCMMVKEGYADGLVSGAIHATADLLRPAFQIIK
                     TAPGAKIVSSFFIMEVPNCEYGENGVFLFADCAVNPSPNAEELASIAVQSANTAKNLLG
                     FEPKVAMLSFSTKGSASHELVDKVRKATEIAKELMPDVAIDGELQLDAALVKEVAELKA
                     PGSKVAGCANVLIFPDLQAGNIGYKLVQRLAKANAIGPITQGMGAPVNDLSRGCSYRDI
                     VDVIATTAVQAQ"
     gene            2273..3331
                     /gene="pta"
                     /label=pta
     CDS             3349..4560
                     /codon_start=1
                     /gene="ack"
                     /product="acetate kinase"
                     /label=ack
                     /note="negative selective marker in Thermoanaerobacter 
                     saccharolyticum"
                     /protein_id="AJP77035.1"
                     /translation="MKIMKILVINCGSSSLKYQLIESTDGNVLAKGLAERIGINDSMLT
                     HNANGEKIKIKKDMKDHKDAIKLVLDALVNSDYGVIKDMSEIDAVGHRVVHGGESFTSS
                     VLINDEVLKAITDCIELAPLHNPANIEGIKACQQIMPNVPMVAVFDTAFHQTMPDYAYL
                     YPIPYEYYTKYRIRRYGFHGTSHKYVSNRAAEILNKPIEDLKIITCHLGNGSSIAAVKY
                     GKSIDTSMGFTPLEGLAMGTRSGSIDPSIISYLMEKENISAEEVVNILNKKSGVYGISG
                     ISSDFRDLEDAAFKNGDERAQLALNVFAYRVKKTIGAYAAAMGGVDVIVFTAGVGENGP
                     EIREFILDGLEFLGFSLDKEKNKVRGKETIISTPNSKVSVMVVPTNEEYMIAKDTEKIV
                     KSIK"
     gene            3349..4560
                     /gene="ack"
                     /label=ack
     regulatory      4688..5218
                     /label=kanamycin promoter
                     /note="kanamycin promoter"
                     /regulatory_class="promoter"
     CDS             5219..6010
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     misc_feature    6349..7260
                     /label=downstream of Tsac_1067
                     /note="downstream of Tsac_1067"
     rep_origin      complement(7670..8215)
                     /direction=LEFT
                     /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."
     promoter        8327..8355
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein
                     gene"
     CDS             complement(8432..9232)
                     /label=URA3
                     /note="orotidine-5'-phosphate decarboxylase, required for
                     uracil biosynthesis"
     promoter        complement(9233..9453)
                     /label=URA3 promoter
     misc_feature    9481..9984
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
ORIGIN
        1 ctagagtcga tcttcgccag cagggcgagg atcgtggcat caccgaaccg cgccgtgcgc
       61 gggtcgtcgg tgagccagag tttcagcagg ccgcccaggc ggcccaggtc gccattgatg
      121 cgggccagct cgcggacgtg ctcatagtcc acgacgcccg tgattttgta gccctggccg
      181 acggccagca ggtaggccga caggctcatg ccggccgccg ccgccttttc ctcaatcgct
      241 cttcgttcgt ctggaaggca gtacaccttg ataggtgggc tgcccttcct ggttggcttg
      301 gtttcatcag ccatccgctt gccctcatct gttacgccgg cggtagccgg ccagcctcgc
      361 agagcaggat tcccgttgag caccgccagg tgcgaataag ggacagtgaa gaaggaacac
      421 ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct gacgccgttg gatacaccaa
      481 ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc gtgcgaaaaa ggatggatat
      541 accgaaaaaa tcgctataat gaccccgaag cagggttatg cagcggaaaa gcgctgcttc
      601 cctgctgttt tgtggaatat ctaccgactg gaaacaggca aatgcaggaa attactgaac
      661 tgaggggaca ggcgagagac gatgccaaag agctacaccg acgagctggc cgagtgggtt
      721 gaatcccgcg cggccaagaa gcgccggcgt gatgaggctg cggttgcgtt cctggcggtg
      781 agggcggatg tcgatatgcg taaggagaaa ataccgcatc aggcgcatat ttgaatgtat
      841 ttagaaaaat aaacaaaaag agtttgtaga aacgcaaaaa ggccatccgt caggatggcc
      901 ttctgcttaa tttgatgcct ggcagtttat ggcgggcgtc ctgcccgcca ccctccgggc
      961 cgttgcttcg caacgttcaa atccgctccc ggcggatttg tcctactcag gagagcgttc
     1021 accgacaaac aacagataaa acgaaaggcc cagtctttcg actgagcctt tcgttttatt
     1081 tgatgcctgg ctcatcgagg tatccaagcg attcaatagt aacagtcctt gtatgccctc
     1141 tttctacagg cacacagaca ttgccagtaa atgtgacgac gaatttgaat ctaaagatag
     1201 tcaaggtaaa tccacaatct gtggacgtca atattcaatg atcttcattt tataaagagg
     1261 atgattaaaa tggattttaa acgacttacc gtagttcttg ggcattatgg cacaggaaaa
     1321 acagaaataa ctataaatta cgcaaggtat ataaagacat gtggatatga tgtagacata
     1381 gtagatcttg atatagtaaa tccatatttt cgcccaagag aagttagaaa agagctttta
     1441 gatggatggg gcataaaagt aattggtgtg gatgaaaagt acatgaatgc tgacatgccg
     1501 gcattatcaa gagatatata cagcgtcata caggatggtg aaaagtatgc tgtatttgat
     1561 gtaggaggcg atgatgcagg agcaatcgca tttgggcaat atcacaatat attagctgat
     1621 aaagatgtag atgtgttttt tgtaataaat gtgaacaggc cgtttacaaa agatgcagaa
     1681 tctgcaataa agtacatgac tcttgtagaa aatgcggcaa gaattaaagt gacacatctt
     1741 gtaaataaca cacatctgtc gtacgagacg atggcagacg atgtaataaa agggcttgaa
     1801 gtgactgaag atgtatcaag gaaaacagga atacccatta agatgacagc cgctaaaaga
     1861 gatgtggctg aaaagcttaa aaatatggga tatgacgttt tccctataga tctttacatg
     1921 aaaacacctt ggaataaaat atgatggaga tgatgaaatt gaagcaagtc atatttaacg
     1981 aaaacttgtg caaaagctgt gaactatgtg ttgaagcatg tccaaaacac ataatagaaa
     2041 tggacaaaga caaactaaac gtaaaaggat accatccagc catgataaaa gaagaaaata
     2101 tggaaaaatg cataagctgc ggataaaccg ctaaggcatg atagctaaag cgtgcccatt
     2161 gtgaagtgga ttgtattcta caattaaacc taatacgctc ataatatgcg cctttctaaa
     2221 aaattattaa ttgtacttat tattttataa aaaatatgtt aaaatgtaaa atgtgtatac
     2281 aatatatttc ttcttagtaa gaggaatgta taaaaataaa tattttaaag gaagggacga
     2341 tcttatgagc attattcaaa acatcattga aaaagctaaa agcgataaaa agaaaattgt
     2401 tctgccagaa ggtgcagaac ccaggacatt aaaagctgct gaaatagttt taaaagaagg
     2461 gattgcagat ttagtgcttc ttggaaatga agatgagata agaaatgctg caaaagactt
     2521 ggacatatcc aaagctgaaa tcattgaccc tgtaaagtct gaaatgtttg ataggtatgc
     2581 taatgatttc tatgagttaa ggaagaacaa aggaatcacg ttggaaaaag ccagagaaac
     2641 aatcaaggat aatatctatt ttggatgtat gatggttaaa gaaggttatg ctgatggatt
     2701 ggtatctggc gctattcatg ctactgcaga tttattaaga cctgcatttc agataattaa
     2761 aacggctcca ggagcaaaga tagtatcaag cttttttata atggaagtgc ctaattgtga
     2821 atatggtgaa aatggtgtat tcttgtttgc tgattgtgcg gtcaacccat cgcctaatgc
     2881 agaagaactt gcttctattg ccgtacaatc tgctaatact gcaaagaatt tgttgggctt
     2941 tgaaccaaaa gttgccatgc tatcattttc tacaaaaggt agtgcatcac atgaattagt
     3001 agataaagta agaaaagcga cagagatagc aaaagaattg atgccagatg ttgctatcga
     3061 cggtgaattg caattggatg ctgctcttgt taaagaagtt gcagagctaa aagcgccggg
     3121 aagcaaagtt gcgggatgtg caaatgtgct tatattccct gatttacaag ctggtaatat
     3181 aggatataag cttgtacaga ggttagctaa ggcaaatgca attggaccta taacacaagg
     3241 aatgggtgca ccggttaatg atttatcaag aggatgcagc tatagagata ttgttgacgt
     3301 aatagcaaca acagctgtgc aggctcaata aaatgtaaag tatggaggat gaaaattatg
     3361 aaaatactgg ttattaattg cggaagttct tcgctaaaat atcaactgat tgaatcaact
     3421 gatggaaatg tgttggcaaa aggccttgct gaaagaatcg gcataaatga ttccatgttg
     3481 acacataatg ctaacggaga aaaaatcaag ataaaaaaag acatgaaaga tcacaaagac
     3541 gcaataaaat tggttttaga tgctttggta aacagtgact acggcgttat aaaagatatg
     3601 tctgagatag atgctgtagg acatagagtt gttcacggag gagaatcttt tacatcatca
     3661 gttctcataa atgatgaagt gttaaaagcg ataacagatt gcatagaatt agctccactg
     3721 cacaatcctg ctaatataga aggaattaaa gcttgccagc aaatcatgcc aaacgttcca
     3781 atggtggcgg tatttgatac agcctttcat cagacaatgc ctgattatgc atatctttat
     3841 ccaatacctt atgaatacta cacaaagtac aggattagaa gatatggatt tcatggcaca
     3901 tcgcataaat atgtttcaaa tagggctgca gagattttga ataaacctat tgaagatttg
     3961 aaaatcataa cttgtcatct tggaaatggc tccagcattg ctgctgtcaa atatggtaaa
     4021 tcaattgaca caagcatggg atttacacca ttagaaggtt tggctatggg tacacgatct
     4081 ggaagcatag acccatccat catttcgtat cttatggaaa aagaaaatat aagcgctgaa
     4141 gaagtagtaa atatattaaa taaaaaatct ggtgtttacg gtatttcagg aataagcagc
     4201 gattttagag acttagaaga tgccgccttt aaaaatggag atgaaagagc tcagttggct
     4261 ttaaatgtgt ttgcatatcg agtaaagaag acgattggcg cttatgcagc agctatggga
     4321 ggcgtcgatg tcattgtatt tacagcaggt gttggtgaaa atggtcctga gatacgagaa
     4381 tttatacttg atggattaga gtttttaggg ttcagcttgg ataaagaaaa aaataaagtc
     4441 agaggaaaag aaactattat atctacgccg aattcaaaag ttagcgtgat ggttgtgcct
     4501 actaatgaag aatacatgat tgctaaagat actgaaaaga ttgtaaagag tataaaatag
     4561 cattcttgac aaatgtttac cccattagta taattaattt tggcaattat attggggtga
     4621 gaaaatgaaa attgatttat caaaaattaa aggacatagg ggccgcagca tcgaagtcaa
     4681 ctacgtaaaa cccagcgaac catttgaggt gataggtaag attataccga ggtatgaaaa
     4741 cgagaattgg acctttacag aattactcta tgaagcgcca tatttaaaaa gctaccaaga
     4801 cgaagaggat gaagaggatg aggaggcaga ttgccttgaa tatattgaca atactgataa
     4861 gataatatat cttttatata gaagatatcg ccgtatgtaa ggatttcagg gggcaaggca
     4921 taggcagcgc gcttatcaat atatctatag aatgggcaaa gcataaaaac ttgcatggac
     4981 taatgcttga aacccaggac aataacctta tagcttgtaa attctatcat aattgtggtt
     5041 tcaaaatcgg ctccgtcgat actatgttat acgccaactt tcaaaacaac tttgaaaaag
     5101 ctgttttctg gtatttaagg ttttagaatg caaggaacag tgaattggag ttcgtcttgt
     5161 tataattagc ttcttggggt atctttaaat actgtagaaa agaggaagga aataataaat
     5221 ggctaaaatg agaatatcac cggaattgaa aaaactgatc gaaaaatacc gctgcgtaaa
     5281 agatacggaa ggaatgtctc ctgctaaggt atataagctg gtgggagaaa atgaaaacct
     5341 atatttaaaa atgacggaca gccggtataa agggaccacc tatgatgtgg aacgggaaaa
     5401 ggacatgatg ctatggctgg aaggaaagct gcctgttcca aaggtcctgc actttgaacg
     5461 gcatgatggc tggagcaatc tgctcatgag tgaggccgat ggcgtccttt gctcggaaga
     5521 gtatgaagat gaacaaagcc ctgaaaagat tatcgagctg tatgcggagt gcatcaggct
     5581 ctttcactcc atcgacatat cggattgtcc ctatacgaat agcttagaca gccgcttagc
     5641 cgaattggat tacttactga ataacgatct ggccgatgtg gattgcgaaa actgggaaga
     5701 agacactcca tttaaagatc cgcgcgagct gtatgatttt ttaaagacgg aaaagcccga
     5761 agaggaactt gtcttttccc acggcgacct gggagacagc aacatctttg tgaaagatgg
     5821 caaagtaagt ggctttattg atcttgggag aagcggcagg gcggacaagt ggtatgacat
     5881 tgccttctgc gtccggtcga tcagggagga tatcggggaa gaacagtatg tcgagctatt
     5941 ttttgactta ctggggatca agcctgattg ggagaaaata aaatattata ttttactgga
     6001 tgaattgttt tagtacctag atttagatgt ctaaaaagct ttttagacat ctaatctttt
     6061 ctgaagtaca tccgcaactg tccatactct gatgttttat atcttttcta aaagttcgct
     6121 agataggggt cccgagcgcc tacgaggaat ttgtatcgga tccgcaagag attatatcga
     6181 gtgcctttaa gaaggctaaa aattacgaag atgtgataca caaaaaggca aaagattacg
     6241 gcaaaaacat accggatagt caagttaaag gagtattgaa acagatagag attactgcct
     6301 taaaccatgt agacaagatt gtcgctgctg aaaagacgat gcagatagag catacgaaag
     6361 aggcaaagaa tttatatctt ctatatcttc tcaacttgtg gcaagataaa acctcccatt
     6421 gtggaggttt catttttaaa gaactttttt tgctgtttta tctattatag ttataggtga
     6481 tatgaatgat tttgtactat tttattaagg ttattacttt agcgattata tccctatttt
     6541 ttggactcaa cgtagattta ttagatgatt tgccggatgg accatgtata tttgtggcta
     6601 atcacaaaag catattagac cctgtagtcc ttatggattc catcgataga agagtttttt
     6661 tcatagccag taaagatttg tataagatac ctattttaaa ttttatttta agcgctttag
     6721 aaacaatacc tatcaaaaaa aattctgcag atgtaaacgc gttaaagaaa gccatcaaaa
     6781 tgcttgatga tggacgatct attgctttgt ttccagaggg aggaatatca cttgacaaaa
     6841 gtgtaataaa aatttacaaa ggtgccatgt atatctcatg caaaacagga tgtccaattg
     6901 tacctgtagg cataagagga actgatggcg tattgcctat gggagagtac tttccacatg
     6961 caggaaagat ttctgtaacg ataggtagaa gcatataccc agatatgtcg ataaataaaa
     7021 atgattgctt ggataagatg gcttatgaag tgttaaattc attaaatagc cttataaata
     7081 atcatgattt aaaatagctg tgatggattg aggtaaatta ttcataggat ttgctcatat
     7141 attaagcata cttgcgttaa attgcaatgg gcttattagt gtgataaaat attatcgtat
     7201 gattttttga tactgaaagg aatggattat atggctagat tatttggcac agatggtgta
     7261 agttaggatg ttggcagacg cttgttgata cgaggcttaa aattgctgag gtaatcggta
     7321 aattcgaccc gtctatcagc tgtccctcct gttcagctac tgacggggtg gtgcgtaacg
     7381 gcaaaagcac cgccggacat cagcgctagc ggagtgtata ctggcttact atgttggcac
     7441 tgatgagggt gtcagtgaag tgcttcatgt ggcaggagaa aaaaggctgc accggtgcgt
     7501 cagcagaata tgtgatacag gatatattcc gcttcctcgc tcactgactc gctacgctcg
     7561 gtcgttcgac tgcggcgagc ggaaatggct tacgaacggg gcggagattt cctggaagat
     7621 gccaggaaga tacttaacag ggaagtgaga gggccgcggc aaagccgttt ttccataggc
     7681 tccgcccccc tgacaagcat cacgaaatct gacgctcaaa tcagtggtgg cgaaacccga
     7741 caggactata aagataccag gcgtttcccc ctggcggctc cctcgtgcgc tctcctgttc
     7801 ctgcctttcg gtttaccggt gtcattccgc tgttatggcc gcgtttgtct cattccacgc
     7861 ctgacactca gttccgggta ggcagttcgc tccaagctgg actgtatgca cgaacccccc
     7921 gttcagtccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggaaaga
     7981 catgcaaaag caccactggc agcagccact ggtaattgat ttagaggagt tagtcttgaa
     8041 gtcatgcgcc ggttaaggct aaactgaaag gacaagtttt ggtgactgcg ctcctccaag
     8101 ccagttacct cggttcaaag agttggtagc tcagagaacc ttcgaaaaac cgccctgcaa
     8161 ggcggttttt tcgttttcag agcaagagat tacgcgcaga ccaaaacgat ctcaagaaga
     8221 tcatcttatt aatcagataa aatatttcta gatttcagtg caatttatct cttcaaatgt
     8281 agcacctgaa gtcagcccca tacgatataa gttgtaattc tcatgtttga cagcttatca
     8341 tcgataagct ttaatgcggt agtttatcac agttaaattg ctaacgcagt caggcaccta
     8401 tacatgcatt tacttataat acagtttttt agttttgctg gccgcatctt ctcaaatatg
     8461 cttcccagcc tgcttttctg taacgttcac cctctacctt agcatccctt ccctttgcaa
     8521 atagtcctct tccaacaata ataatgtcag atcctgtaga gaccacatca tccacggttc
     8581 tatactgttg acccaatgcg tctcccttgt catctaaacc cacaccgggt gtcataatca
     8641 accaatcgta accttcatct cttccaccca tgtctctttg agcaataaag ccgataacaa
     8701 aatctttgtc gctcttcgca atgtcaacag tacccttagt atattctcca gtagataggg
     8761 agcccttgca tgacaattct gctaacatca aaaggcctct aggttccttt gttacttctt
     8821 ctgccgcctg cttcaaaccg ctaacaatac ctgggcccac cacaccgtgt gcattcgtaa
     8881 tgtctgccca ttctgctatt ctgtatacac ccgcagagta ctgcaatttg actgtattac
     8941 caatgtcagc aaattttctg tcttcgaaga gtaaaaaatt gtacttggcg gataatgcct
     9001 ttagcggctt aactgtgccc tccatggaaa aatcagtcaa gatatccaca tgtgttttta
     9061 gtaaacaaat tttgggacct aatgcttcaa ctaactccag taattccttg gtggtacgaa
     9121 catccaatga agcacacaag tttgtttgct tttcgtgcat gatattaaat agcttggcag
     9181 caacaggact aggatgagta gcagcacgtt ccttatatgt agctttcgac atgatttatc
     9241 ttcgtttcct gcaggttttt gttctgtgca gttgggttaa gaatactggg caatttcatg
     9301 tttcttcaac actacatatg cgtatatata ccaatctaag tctgtgctcc ttccttcgtt
     9361 cttccttctg ttcggagatt accgaatcaa aaaaatttca aagaaaccga aatcaaaaaa
     9421 aagaataaaa aaaaaatgat gaattgaatt gaaaagctag cttatcgatg ggtccttttc
     9481 atcacgtgct ataaaaataa ttataattta aattttttaa tataaatata taaattaaaa
     9541 atagaaagta aaaaaagaaa ttaaagaaaa aatagttttt gttttccgaa gatgtaaaag
     9601 actctagggg gatcgccaac aaatactacc ttttatcttg ctcttcctgc tctcaggtat
     9661 taatgccgaa ttgtttcatc ttgtctgtgt agaagaccac acacgaaaat cctgtgattt
     9721 tacattttac ttatcgttaa tcgaatgtat atctatttaa tctgcttttc ttgtctaata
     9781 aatatatatg taaagtacgc tttttgttga aattttttaa acctttgttt attttttttt
     9841 cttcattccg taactcttct accttcttta tttactttct aaaatccaaa tacaaaacat
     9901 aaaaataaat aaacacagag taaattccca aattattcca tcattaaaag atacgaggcg
     9961 cgtgtaagtt acaggcaagc gatctctaag aaaccattat tatcatgaca ttaacctata
    10021 aaaaaggcct ctcgag
//
