LOCUS       V008287                 8237 bp    DNA     circular SYN 17-DEC-2018
DEFINITION  Exported.
ACCESSION   V008287
VERSION     V008287
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
REFERENCE   1  (bases 1 to 8237)
  AUTHORS   Chandran S, Noskov VN, Segall-Shapiro TH, Ma L, Whiteis C, Lartigue
            C, Jores J, Vashee S, Chuang R.
  TITLE     TREC-IN: gene knock-in genetic tool for genomes cloned in yeast
  JOURNAL   BMC Genomics 15 (1), 1180 (2014)
   PUBMED   25539750
REFERENCE   2  (bases 1 to 8237)
  AUTHORS   Chuang R-Y., Noskov V, Ma L.
  TITLE     Direct Submission
  JOURNAL   Submitted (17-DEC-2014) Synthetic Biology, J. Craig Venter
            Institute, 9704 Medical Center Drive, Rockville, MD 20850, USA
REFERENCE   3  (bases 1 to 8237)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8237)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing
            ##Assembly-Data-END##
            SGRef: number: 1; type: "Journal Article"; journalName: "BMC
            Genomics"; date: "2014"; volume: "15"; issue: "1"; pages: "1180"
            SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
            (17-DEC-2014) Synthetic Biology, J. Craig Venter Institute, 9704
            Medical Center Drive, Rockville, MD 20850, USA"
            SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8237
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        315..502
                     /label="HIS3 promoter"
     CDS             503..1159
                     /label="HIS3"
                     /note="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
     rep_origin      complement(1423..1878)
                     /direction=LEFT
                     /label="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     2023..2039
                     /label="M13 fwd"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        2046..2064
                     /label="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2097..2113
                     /label="SK primer"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     misc_feature    2118..2131
                     /label="I-SceI restriction site"
                     /note="I-SceI restriction site"
     promoter        2133..2574
                     /label="GAL1 promoter"
                     /note="inducible promoter, regulated by Gal4"
     CDS             2597..3301
                     /gene="SCEI"
                     /label="Intron-encoded endonuclease I-SceI"
                     /note="Intron-encoded endonuclease I-SceI from
                     Saccharomyces cerevisiae (strain ATCC 204508 / S288c).
                     Accession#: P03882"
     CDS             complement(3379..4182)
                     /codon_start=1
                     /gene="ura3"
                     /product="Ura3"
                     /label="ura3"
                     /protein_id="AJD09819.1"
                     /translation="MSTKSYTSRAETHASPVASKLLRLMDEKKTNLCASLDVRSTDELL
                     KLVETLGPYICLLKTHVDILDDFSYEGTVVPLKALAEKYKFLIFEDRKFADIGNTVKLQ
                     YTSGVYRIAEWSDITNAHGVTGAGIVAGLKQGAQEVTKEPRGLLMLAELSSKGSLAHGE
                     YTKGTVDIAKSDKDFVIGFIAQNDMGGREEGFDWLIMTPGVGLDDKGDALGQQYRTVDE
                     VVSGGSDIIIVGRGLFAKGRDPKVEGERYRNAGWEAYQKRISAPH"
     gene            complement(3379..4182)
                     /gene="ura3"
                     /label="ura3"
                     /note="KlUra3"
     promoter        4498..4841
                     /label="TEF promoter"
                     /note="Ashbya gossypii TEF promoter"
     CDS             4882..5396
                     /codon_start=1
                     /gene="kanMX4"
                     /product="kanamycin resistance protein"
                     /label="kanMX4"
                     /note="KanR"
                     /protein_id="AJD09820.1"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGW"
     gene            4882..5396
                     /gene="kanMX4"
                     /label="kanMX4"
     primer_bind     complement(5417..5433)
                     /label="KS primer"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     promoter        complement(5463..5481)
                     /label="T3 promoter"
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5502..5518)
                     /label="M13 rev"
                     /note="common sequencing primer, one of multiple similar
                     variants"
     protein_bind    complement(5526..5542)
                     /label="lac operator"
                     /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)."
     promoter        complement(5550..5580)
                     /label="lac promoter"
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5595..5616)
                     /label="CAP binding site"
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5904..6492)
                     /direction=LEFT
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
     CDS             complement(6666..7523)
                     /label="AmpR"
                     /note="beta-lactamase"
     promoter        complement(7524..7628)
                     /label="AmpR promoter"
     misc_feature    7665..8168
                     /label="CEN/ARS"
                     /note="S. cerevisiae CEN6 centromere fused to an
                     autonomously replicating sequence"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accataattc cgttttaaga gcttggtgag cgctaggagt cactgccagg tatcgtttga
      241 acacggcatt agtcagggaa gtcataacac agtcctttcc cgcaattttc tttttctatt
      301 actcttggcc tcctctagta cactctatat ttttttatgc ctcggtaatg attttcattt
      361 ttttttttcc acctagcgga tgactctttt tttttcttag cgattggcat tatcacataa
      421 tgaattatac attatataaa gtaatgtgat ttcttcgaag aatatactaa aaaatgagca
      481 ggcaagataa acgaaggcaa agatgacaga gcagaaagcc ctagtaaagc gtattacaaa
      541 tgaaaccaag attcagattg cgatctcttt aaagggtggt cccctagcga tagagcactc
      601 gatcttccca gaaaaagagg cagaagcagt agcagaacag gccacacaat cgcaagtgat
      661 taacgtccac acaggtatag ggtttctgga ccatatgata catgctctgg ccaagcattc
      721 cggctggtcg ctaatcgttg agtgcattgg tgacttacac atagacgacc atcacaccac
      781 tgaagactgc gggattgctc tcggtcaagc ttttaaagag gccctactgg cgcgtggagt
      841 aaaaaggttt ggatcaggat ttgcgccttt ggatgaggca ctttccagag cggtggtaga
      901 tctttcgaac aggccgtacg cagttgtcga acttggtttg caaagggaga aagtaggaga
      961 tctctcttgc gagatgatcc cgcattttct tgaaagcttt gcagaggcta gcagaattac
     1021 cctccacgtt gattgtctgc gaggcaagaa tgatcatcac cgtagtgaga gtgcgttcaa
     1081 ggctcttgcg gttgccataa gagaagccac ctcgcccaat ggtaccaacg atgttccctc
     1141 caccaaaggt gttcttatgt agtgacaccg attatttaaa gctgcagcat acgatatata
     1201 tacatgtgta tatatgtata cctatgaatg tcagtaagta tgtatacgaa cagtatgata
     1261 ctgaagatga caaggtaatg catcattcta tacgtgtcat tctgaacgag gcgcgctttc
     1321 cttttttctt tttgcttttt cttttttttt ctcttgaact cgacggatca tatgcggtgt
     1381 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggaaattgta aacgttaata
     1441 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     1501 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     1561 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     1621 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     1681 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     1741 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     1801 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     1861 cgccgctaca gggcgcgtcg cgccattcgc cattcaggct gcgcaactgt tgggaagggc
     1921 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa ggggggatgt gctgcaaggc
     1981 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     2041 aattgtaata cgactcacta tagggcgaat tggagctcca ccgcggtggc ggccgctcta
     2101 gaactagtgg atcccccgat aacagggtaa tacggattag aagccgccga gcgggtgaca
     2161 gccctccgaa ggaagactct cctccgtgcg tcctcgtctt caccggtcgc gttcctgaaa
     2221 cgcagatgtg cctcgcgccg cactgctccg aacaataaag attctacaat actagctttt
     2281 atggttatga agaggaaaaa ttggcagtaa cctggcccca caaaccttca aatgaacgaa
     2341 tcaaattaac aaccatagga tgataatgcg attagttttt tagccttatt tctggggtaa
     2401 ttaatcagcg aagcgatgat ttttgatcta ttaacagata tataaatgca aaaactgcat
     2461 aaccacttta actgatactt tcaacatttt cggtttgtat tacttcttat tcaaatgtaa
     2521 taaaagtatc aacaaaaaat tgttaatata cctctatact ttaacgtcaa ggagaaaaaa
     2581 ccccggatcc atgcatatga aaaacatcaa aaaaaaccag gtaatgaacc tgggtccgaa
     2641 ctctaaactg ctgaaagaat acaaatccca gctgatcgaa ctgaacatcg aacagttcga
     2701 agcaggtatc ggtctgatcc tgggtgatgc ttacatccgt tctcgtgatg aaggtaaaac
     2761 ctactgtatg cagttcgagt ggaaaaacaa agcatacatg gaccacgtat gtctgctgta
     2821 cgatcagtgg gtactgtccc cgccgcacaa aaaagaacgt gttaaccacc tgggtaacct
     2881 ggtaatcacc tggggcgccc agactttcaa acaccaagct ttcaacaaac tggctaacct
     2941 gttcatcgtt aacaacaaaa aaaccatccc gaacaacctg gttgaaaact acctgacccc
     3001 gatgtctctg gcatactggt tcatggatga tggtggtaaa tgggattaca acaaaaactc
     3061 taccaacaaa tcgatcgtac tgaacaccca gtctttcact ttcgaagaag tagaatacct
     3121 ggttaagggt ctgcgtaaca aattccaact gaactgttac gtaaaaatca acaaaaacaa
     3181 accgatcatc tacatcgatt ctatgtctta cctgatcttc tacaacctga tcaaaccgta
     3241 cctgatcccg cagatgatgt acaaactgcc gaacactatc tcctccgaaa ctttcctgaa
     3301 ataagagtgt tgcaccgtgc caatgcaggt ggctattaga ttaaatatgt gatttgttct
     3361 attaagtttc ctgtataatt aatggggagc gctgattctc ttttggtacg cttcccatcc
     3421 agcatttctg tatctttcac cttcaacctt aggatctcta cccttggcga aaagtcctct
     3481 gccaacaatg atgatatctg atccaccact tacaacttcg tcgacggttc tgtactgctg
     3541 acccaatgca tcgcctttgt cgtctaaacc tacacctggg gtcatgatta gccaatcaaa
     3601 cccttcttct cttcctccca tatcgttctg agcaatgaac ccaataacga aatctttatc
     3661 actctttgca atatcaacgg tacccttagt atattcaccg tgtgctagag aacccttgga
     3721 agacaattca gcaagcatca ataatcccct tggttctttg gtgacctctt gcgcaccttg
     3781 tttcaagcca gcaacaatac cagcaccagt aaccccgtgg gcgttggtga tatcagacca
     3841 ttctgcgata cggtaaacgc ccgatgtata ttgtaatttg actgtgttac cgatatcggc
     3901 gaattttctg tcctcaaata tcaagaactt gtatttctct gccaatgctt tcaatggaac
     3961 gacagtaccc tcataactga aatcatccaa gatatcaacg tgtgttttca aaaggcaaat
     4021 gtatggaccc aacgtttcaa ctagtttcaa tagctcatca gtcgaacgaa cgtcaagaga
     4081 agcacacaaa ttggtcttct tttcatccat taaacgtaaa agtttcgatg caaccggact
     4141 tgcatgagtc tcagctctac tggtatatga ttttgtggac atggtgcaac taattgacgg
     4201 gagtgtattg acgctggcgt actggctttc acaaaatggc ccaatcacaa ccacatctta
     4261 gatagttgaa atgactttag ataacatcaa ttgagatgag cttaatcatg tcaaagctaa
     4321 aagtgtcacc atgaacgaca attcttaagc aaatcacgtg atatagatcc acgaataacc
     4381 accatttgat gctcgaggca agtaatgtgt gtaaaaaaat gcgttaccac catccaatgc
     4441 agaccgatct tctacccaga atcacatata tttatgtacc gagggctgca ggaattcgac
     4501 atggaggccc agaataccct ccttgacagt cttgacgtgc gcagctcagg ggcatgatgt
     4561 gactgtcgcc cgtacattta gcccatacat ccccatgtat aatcatttgc atccatacat
     4621 tttgatggcc gcacggcgcg aagcaaaaat tacggctcct cgctgcagac ctgcgagcag
     4681 ggaaacgctc ccctcacaga cgcgttgaat tgtccccacg ccgcgcccct gtagagaaat
     4741 ataaaaggtt aggatttgcc actgaggttc ttctttcata tacttccttt taaaatcttg
     4801 ctaggataca gttctcacat cacatccgaa cataaacaac cactagttac cgttcgtata
     4861 agaaaccata tacgaagtta tatgggtaag gaaaagactc acgtttcgag gccgcgatta
     4921 aattccaaca tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa
     4981 tcaggtgcga caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa
     5041 catggcaaag gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg
     5101 acggaattta tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg
     5161 ttactcacca ctgcgatccc cggcaaaaca gcattccagg tattagaaga atatcctgat
     5221 tcaggtgaaa atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct
     5281 gtttgtaatt gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga
     5341 atgaataacg gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggccgaat
     5401 tcgatatcaa gcttatcgat accgtcgacc tcgagggggg gcccggtacc cagcttttgt
     5461 tccctttagt gagggttaat tccgagcttg gcgtaatcat ggtcatagct gtttcctgtg
     5521 tgaaattgtt atccgctcac aattccacac aacataggag ccggaagcat aaagtgtaaa
     5581 gcctggggtg cctaatgagt gaggtaactc acattaattg cgttgcgctc actgcccgct
     5641 ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga
     5701 ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     5761 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     5821 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     5881 aaaaaggccg cgttgctggc gtttttccat aggctcggcc cccctgacga gcatcacaaa
     5941 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttc
     6001 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6061 tccgcctttc tcccttcggg aagcgtggcg ctttctcaat gctcacgctg taggtatctc
     6121 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6181 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6241 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6301 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     6361 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6421 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     6481 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     6541 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     6601 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     6661 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6721 atagttgcct gactgcccgt cgtgtagata actacgatac gggagggctt accatctggc
     6781 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     6841 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6901 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6961 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7021 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgaaaaaaa
     7081 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7141 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7201 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7261 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7321 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7381 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7441 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7501 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     7561 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     7621 gttccgcgca catttccccg aaaagtgcca cctgggtcct tttcatcacg tgctataaaa
     7681 ataattataa tttaaatttt ttaatataaa tatataaatt aaaaatagaa agtaaaaaaa
     7741 gaaattaaag aaaaaatagt ttttgttttc cgaagatgta aaagactcta gggggatcgc
     7801 caacaaatac taccttttat cttgctcttc ctgctctcag gtattaatgc cgaattgttt
     7861 catcttgtct gtgtagaaga ccacacacga aaatcctgtg attttacatt ttacttatcg
     7921 ttaatcgaat gtatatctat ttaatctgct tttcttgtct aataaatata tatgtaaagt
     7981 acgctttttg ttgaaatttt ttaaaccttt gtttattttt ttttcttcat tccgtaactc
     8041 ttctaccttc tttatttact ttctaaaatc caaatacaaa acataaaaat aaataaacac
     8101 agagtaaatt cccaaattat tccatcatta aaagatacga ggcgcgtgta agttacaggc
     8161 aagcgatccg tcctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     8221 cacgaggccc tttcgtc
//
