LOCUS Exported File 6499 bp ds-DNA circular SYN 30-JAN-2012 DEFINITION Addgene ID 34973. ACCESSION . VERSION . KEYWORDS pDS277 SOURCE synthetic DNA construct ORGANISM recombinant plasmid REFERENCE 1 (bases 1 to 6499) AUTHORS Devin Strickland TITLE Direct Submission JOURNAL Exported Jan 30, 2012 from SnapGene 0.6.2.2 http://www.snapgene.com COMMENT TULIPs cassette is inserted into YIPlac128 between SacI and SphI. SphI was destroyed. FEATURES Location/Qualifiers source 1..6499 /organism="recombinant plasmid" /lab_host="Saccharomyces cerevisiae" /mol_type="other DNA" promoter 22..463 /gene="S. cerevisiae GAL1" /note="GAL1 promoter" /note="inducible promoter, regulated by Gal4" /note="color: #c0c0c0; direction: RIGHT" CDS 549..776 /codon_start=1 /note="Mid2-SS/TM" /note="color: #cc99b2" /translation="MLSFTTKNSFRLLLLILSCISTIRAQFFVQSSSSNSGLSKKNRNI VIGCVVGIGVPLILVILALIYMFCIQSSRTD" CDS 792..1505 /codon_start=1 /note="GFP(S65T)" /note="color: #00ff00" /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTFTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" CDS 1539..1973 /codon_start=1 /allele="V529N" /note="LOVpep" /note="Amino acids numbered according to A. sativa phototropin 1" /note="color: #666699" /translation="LATTLERIEKNFVITDPRLPDNPIIFASDSFLQLTEYSREEILGR NCRFLQGPETDRATVRKIRDAIDNQTEVTVQLINYTKSGKKFWNLFHLQPMRDQKGDVQ YFIGVQLDGTEHVRDAAEREANMLIKKTAEEIDKAVDTWV" terminator 1984..2231 /note="CYC1 term" /note="color: #c0c0c0" primer_bind complement(2256..2272) /note="M13 rev" /note="common sequencing primer, one of multiple similar variants" /note="color: #a020f0; direction: LEFT" rep_origin complement(2658..3246) /direction=LEFT /note="ori" /note="high-copy-number colE1/pMB1/pBR322/pUC origin of replication" /note="color: #ffff00" CDS complement(3417..4277) /codon_start=1 /gene="bla" /product="beta-lactamase" /note="AmpR" /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /note="This feature has 2 segments: 1: 3417 .. 4208 / #ccffcc 2: 4209 .. 4277 / #ccffcc / signal sequence Cleavage site after base 4208" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" promoter complement(4278..4382) /gene="bla" /note="AmpR promoter" /note="color: #ffffff; direction: LEFT" promoter 4470..4874 /gene="S. cerevisiae LEU2" /note="LEU2 promoter" /note="color: #ffffff; direction: RIGHT" CDS 4887..5981 /codon_start=1 /gene="S. cerevisiae LEU2" /product="3-isopropylmalate dehydrogenase, required for leucine biosynthesis" /note="LEU2" /note="yeast auxotrophic marker" /note="color: #ff7f50" /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG DLGGSNSTTEVGDAVAEEVKKILA" primer_bind 6472..6488 /note="M13 fwd" /note="common sequencing primer, one of multiple similar variants" /note="color: #a020f0; direction: RIGHT" ORIGIN 1 cgtttaacgg cgcgcctagt acggattaga agccgccgag cgggtgacag ccctccgaag 61 gaagactctc ctccgtgcgt cctcgtcttc accggtcgcg ttcctgaaac gcagatgtgc 121 ctcgcgccgc actgctccga acaataaaga ttctacaata ctagctttta tggttatgaa 181 gaggaaaaat tggcagtaac ctggccccac aaaccttcaa atgaacgaat caaattaaca 241 accataggat gataatgcga ttagtttttt agccttattt ctggggtaat taatcagcga 301 agcgatgatt tttgatctat taacagatat ataaatgcaa aaactgcata accactttaa 361 ctaatacttt caacattttc ggtttgtatt acttcttatt caaatgtaat aaaagtatca 421 acaaaaaatt gttaatatac ctctatactt taacgtcaag gagaaaaacc ccggattctg 481 aagcggccgc actagtggat cccccgggct gcaggaattc gatatcaagc ttatcgatac 541 cgtcgacaat gttgtctttc acaaccaaga atagtttccg cttattactt ttaatactgt 601 catgcatatc gacgatacgc gcacaatttt tcgtgcaatc atcatcttcg aattcgggtc 661 tttctaaaaa aaatagaaac atcgtcattg gttgtgtggt tggtattggt gtaccactga 721 ttctggtgat actggcttta atttacatgt tttgtatcca atcatcaagg acggacgcgt 781 cgacctcgac aatgagtaaa ggagaagaac ttttcactgg agttgtccca attcttgttg 841 aattagatgg tgatgttaat gggcacaaat tttctgtcag tggagagggt gaaggtgatg 901 caacatacgg aaaacttacc cttaaattta tttgcactac tggaaaacta cctgttccat 961 ggccaacact tgtcactact ttcacttatg gtgttcaatg cttttcaaga tacccagatc 1021 atatgaaaca gcatgacttt ttcaagagtg ccatgcccga aggttatgta caggaaagaa 1081 ctatattttt caaagatgac gggaactaca agacacgtgc tgaagtcaag tttgaaggtg 1141 atacccttgt taatagaatc gagttaaaag gtattgattt taaagaagat ggaaacattc 1201 ttggacacaa attggaatac aactataact cacacaatgt atacatcatg gcagacaaac 1261 aaaagaatgg aatcaaagtt aacttcaaaa ttagacacaa cattgaagat ggaagcgttc 1321 aactagcaga ccattatcaa caaaatactc caattggcga tggccctgtc cttttaccag 1381 acaaccatta cctgtccaca caatctgccc tttcgaaaga tcccaacgaa aagagagacc 1441 acatggtcct tcttgagttt gtaacagctg ctgggattac acatggcatg gatgaactat 1501 acaaaggtgg atctggaggt tcaggtggaa gtcctaggtt ggctactaca cttgaacgta 1561 ttgagaagaa ctttgtcatt actgacccaa gattgccaga taatcccatt atattcgcgt 1621 ccgatagttt cttgcagttg acagaatata gccgtgaaga aattttggga agaaactgca 1681 ggtttctaca aggtcctgaa actgatcgcg cgacagtgag aaaaattaga gatgccatag 1741 ataaccaaac agaggtcact gttcagctga ttaattatac aaagagtggt aaaaagttct 1801 ggaacctctt tcacttgcag cctatgcgag atcagaaggg agatgtccag tactttattg 1861 gggttcagtt ggatggaact gagcatgtcc gagatgctgc cgagagagag gcaaacatgc 1921 tgattaagaa aactgcagaa gagattgata aagcggtgga tacctgggtg taagcttctc 1981 gagtcatgta attagttatg tcacgcttac attcacgccc tccccccaca tccgctctaa 2041 ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt tttatagtta 2101 tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg tacagacgcg 2161 tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg 2221 ctttaatttg cggccgcaag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat 2281 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg 2341 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag 2401 tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt 2461 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg 2521 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg 2581 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag 2641 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga 2701 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct 2761 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc 2821 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg 2881 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc 2941 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca 3001 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag 3061 ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct 3121 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc 3181 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga 3241 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca 3301 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat 3361 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac 3421 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt 3481 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt 3541 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag 3601 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct 3661 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt 3721 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc 3781 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt 3841 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg 3901 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg 3961 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct 4021 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc 4081 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt 4141 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt 4201 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg 4261 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat 4321 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg 4381 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta 4441 acctataaaa ataggcgtat cacgaggcca actgtgggaa tactcaggta tcgtaagatg 4501 caagagttcg aatctcttag caaccattat ttttttcctc aacataacga gaacacacag 4561 gggcgctatc gcacagaatc aaattcgatg actggaaatt ttttgttaat ttcagaggtc 4621 gcctgacgca tatacctttt tcaactgaaa aattgggaga aaaaggaaag gtgagaggcc 4681 ggaaccggct tttcatatag aatagagaag cgttcatgac taaatgcttg catcacaata 4741 cttgaagttg acaatattat ttaaggacct attgtttttt ccaataggtg gttagcaatc 4801 gtcttacttt ctaacttttc ttacctttta catttcagca atatatatat atatttcaag 4861 gatataccat tctaatgtct gcccctatgt ctgcccctaa gaagatcgtc gttttgccag 4921 gtgaccacgt tggtcaagaa atcacagccg aagccattaa ggttcttaaa gctatttctg 4981 atgttcgttc caatgtcaag ttcgatttcg aaaatcattt aattggtggt gctgctatcg 5041 atgctacagg tgtcccactt ccagatgagg cgctggaagc ctccaagaag gttgatgccg 5101 ttttgttagg tgctgtgggt ggtcctaaat ggggtacagg tagtgttaga cctgaacaag 5161 gtttactaaa aatccgtaaa gaacttcaat tgtacgccaa cttaagacca tgtaactttg 5221 catccgactc tcttttagac ttatctccaa tcaagccaca atttgctaaa ggtactgact 5281 tcgttgttgt cagagaatta gtgggaggta tttactttgg taagagaaag gaagacgatg 5341 gtgatggtgt cgcttgggat agtgaacaat acaccgttcc agaagtgcaa agaatcacaa 5401 gaatggccgc tttcatggcc ctacaacatg agccaccatt gcctatttgg tccttggata 5461 aagctaatgt tttggcctct tcaagattat ggagaaaaac tgtggaggaa accatcaaga 5521 acgaatttcc tacattgaag gttcaacatc aattgattga ttctgccgcc atgatcctag 5581 ttaagaaccc aacccaccta aatggtatta taatcaccag caacatgttt ggtgatatca 5641 tctccgatga agcctccgtt atcccaggtt ccttgggttt gttgccatct gcgtccttgg 5701 cctctttgcc agacaagaac accgcatttg gtttgtacga accatgccac ggttctgctc 5761 cagatttgcc aaagaataag gttgacccta tcgccactat cttgtctgct gcaatgatgt 5821 tgaaattgtc attgaacttg cctgaagaag gtaaggccat tgaagatgca gttaaaaagg 5881 ttttggatgc aggtatcaga actggtgatt taggtggttc caacagtacc accgaagtcg 5941 gtgatgctgt cgccgaagaa gttaagaaaa tccttgctta aaaagattct ctttttttat 6001 gatatttgta cataaacttt ataaatgaaa ttcataatag aaacgacacg aaattacaaa 6061 atggaatatg ttcatagggt aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa 6121 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga 6181 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact 6241 atgcggcatc agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca 6301 gatgcgtaag gagaaaatac cgcatcaggc gccattcgcc attcaggctg cgcaactgtt 6361 gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg 6421 ctgcaaggcg attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga 6481 cggccagtga attcgagct //