LOCUS Exported 8218 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION A dual reporter vector modified from Sharon et al (Nature Biotech, 2012) containing Ura3 and Nat1 selectable markers, a constant background RFP (TEF2::mCherry), and a variable YFP (pTpA+MCS::yEVenus).. ACCESSION . VERSION . KEYWORDS yeast_DualReporter SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8218) AUTHORS Carl G. de Boer, Eeshit Dhaval Vaishnav, Ronen Sadeh, Esteban Luis Abeyta, Nir Friedman, Aviv Regev TITLE Deciphering eukaryotic cis-regulatory logic with 100 million random promoters JOURNAL BioRxiv REFERENCE 2 (bases 1 to 8218) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8218 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(93..115) /label=pGEX 3' /note="pGEX vectors, reverse primer" promoter 315..333 /label=SP6 promoter /note="promoter for bacteriophage SP6 RNA polymerase" primer_bind 315..332 /label=SP6 /note="SP6 promoter, forward primer" promoter 407..622 /gene="S. cerevisiae URA3" /label=URA3 promoter CDS 623..1426 /codon_start=1 /gene="S. cerevisiae URA3" /product="orotidine-5'-phosphate decarboxylase, required for uracil biosynthesis" /label=URA3 /note="yeast auxotrophic marker, counterselectable with 5-fluoroorotic acid (5-FOA)" /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN" primer_bind complement(1694..1715) /label=F1ori-F /note="F1 origin, forward primer" CDS 1960..2676 /codon_start=1 /product="yeast-enhanced green fluorescent protein" /label=yeGFP /note="codons optimized for translation in S. cerevisiae and C. albicans" /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK LICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" terminator 2697..2884 /gene="S. cerevisiae ADH1" /label=ADH1 terminator /note="transcription terminator for the S. cerevisiae alcohol dehydrogenase 1 (ADH1) gene" gene 2959..4078 /label=natMX6 /note="yeast selectable marker conferring nourseothricin resistance" promoter 2959..3302 /label=TEF promoter /note="Ashbya gossypii TEF promoter" CDS 3303..3875 /codon_start=1 /gene="Streptomyces noursei nat1" /product="nourseothricin acetyltransferase" /label=NrsR /note="confers resistance to nourseothricin" /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT LCGLDTALYDGTASDGEQALYMSMPCP" terminator 3881..4078 /label=TEF terminator /note="Ashbya gossypii TEF terminator" misc_feature 4145..4648 /label=CEN/ARS /note="S. cerevisiae CEN6 centromere fused to an autonomously replicating sequence" primer_bind complement(4704..4723) /label=T7 /note="T7 promoter, forward primer" promoter complement(4705..4723) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(4810..4827) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4981..5569) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5061..5080) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5740..6600) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 6363..6382 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6601..6705) /gene="bla" /label=AmpR promoter terminator 6770..6957 /gene="S. cerevisiae ADH1" /label=ADH1 terminator /note="transcription terminator for the S. cerevisiae alcohol dehydrogenase 1 (ADH1) gene" CDS complement(6984..7694) /codon_start=1 /product="monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)" /label=mCherry /note="mammalian codon-optimized" /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA EGRHSTGGMDELYK" primer_bind complement(7262..7281) /label=mCherry-F /note="mCherry, forward primer" primer_bind 7531..7549 /label=mCherry-R /note="mCherry, reverse primer" ORIGIN 1 gaagcgccta cgcttgacat ctactatatg taagtatacg gccccatata taggcccttt 61 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg 121 gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg 181 ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga 241 gtgcaccata tggacatatt gtcgttagaa cgcggctaca attaatacat aaccttatgt 301 atcatacaca tacgatttag gtgacactat agaacgcggc cgccagctga agctttaact 361 atgcggcatc agagcagatt gtactgagag tgcaccatac caccttttca attcatcatt 421 ttttttttat tctttttttt gatttcggtt tccttgaaat ttttttgatt cggtaatctc 481 cgaacagaag gaagaacgaa ggaaggagca cagacttaga ttggtatata tacgcatatg 541 tagtgttgaa gaaacatgaa attgcccagt attcttaacc caactgcaca gaacaaaaac 601 ctgcaggaaa cgaagataaa tcatgtcgaa agctacatat aaggaacgtg ctgctactca 661 tcctagtcct gttgctgcca agctatttaa tatcatgcac gaaaagcaaa caaacttgtg 721 tgcttcattg gatgttcgta ccaccaagga attactggag ttagttgaag cattaggtcc 781 caaaatttgt ttactaaaaa cacatgtgga tatcttgact gatttttcca tggagggcac 841 agttaagccg ctaaaggcat tatccgccaa gtacaatttt ttactcttcg aagacagaaa 901 atttgctgac attggtaata cagtcaaatt gcagtactct gcgggtgtat acagaatagc 961 agaatgggca gacattacga atgcacacgg tgtggtgggc ccaggtattg ttagcggttt 1021 gaagcaggcg gcagaagaag taacaaagga acctagaggc cttttgatgt tagcagaatt 1081 gtcatgcaag ggctccctat ctactggaga atatactaag ggtactgttg acattgcgaa 1141 gagcgacaaa gattttgtta tcggctttat tgctcaaaga gacatgggtg gaagagatga 1201 aggttacgat tggttgatta tgacacccgg tgtgggttta gatgacaagg gagacgcatt 1261 gggtcaacag tatagaaccg tggatgatgt ggtctctaca ggatctgaca ttattattgt 1321 tggaagagga ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa 1381 agcaggctgg gaagcatatt tgagaagatg cggccagcaa aactaaaaaa ctgtattata 1441 agtaaatgca tgtatactaa actcacaaat tagagcttca atttaattat atcagttatt 1501 accctatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggaaat 1561 tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt 1621 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg 1681 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt 1741 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc 1801 aagtgctagc aggaatgatg caaaaggttc ccgattcgaa ctgcattttt ttcacatctc 1861 gaggttacgg ctgtttctta attaaaaaaa gatagaaaac attaggagtg taacacaaga 1921 ctttcggatc ctgagcaggc aagataaacg aaggcaaaga tgtctaaagg tgaagaatta 1981 ttcactggtg ttgtcccaat tttggttgaa ttagatggtg atgttaatgg tcacaaattt 2041 tctgtctccg gtgaaggtga aggtgatgct acttacggta aattgacctt aaaattgatt 2101 tgtactactg gtaaattgcc agttccatgg ccaaccttag tcactacttt aggttatggt 2161 ttgcaatgtt ttgctagata cccagatcat atgaaacaac atgacttttt caagtctgcc 2221 atgccagaag gttatgttca agaaagaact atttttttca aagatgacgg taactacaag 2281 accagagctg aagtcaagtt tgaaggtgat accttagtta atagaatcga attaaaaggt 2341 attgatttta aagaagatgg taacatttta ggtcacaaat tggaatacaa ctataactct 2401 cacaatgttt acatcactgc tgacaaacaa aagaatggta tcaaagctaa cttcaaaatt 2461 agacacaaca ttgaagatgg tggtgttcaa ttagctgacc attatcaaca aaatactcca 2521 attggtgatg gtccagtctt gttaccagac aaccattact tatcctatca atctgcctta 2581 tccaaagatc caaacgaaaa gagagaccac atggtcttgt tagaatttgt tactgctgct 2641 ggtattaccc atggtatgga tgaattgtac aaataaggcg cgccacttct aaataagcga 2701 atttcttatg atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca 2761 aattttaaag tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc 2821 tgtaggtcag gttgctttct caggtatagt atgaggtcgc tcttattgac cacacctcta 2881 ccggcagatc cgctagggat aacagggtaa tatagatctg tttagcttgc ctcgtccccg 2941 ccgggtcacc cggccagcga catggaggcc cagaataccc tccttgacag tcttgacgtg 3001 cgcagctcag gggcatgatg tgactgtcgc ccgtacattt agcccataca tccccatgta 3061 taatcatttg catccataca ttttgatggc cgcacggcgc gaagcaaaaa ttacggctcc 3121 tcgctgcaga cctgcgagca gggaaacgct cccctcacag acgcgttgaa ttgtccccac 3181 gccgcgcccc tgtagagaaa tataaaaggt taggatttgc cactgaggtt cttctttcat 3241 atacttcctt ttaaaatctt gctaggatac agttctcaca tcacatccga acataaacaa 3301 ccatgggtac cactcttgac gacacggctt accggtaccg caccagtgtc ccgggggacg 3361 ccgaggccat cgaggcactg gatgggtcct tcaccaccga caccgtcttc cgcgtcaccg 3421 ccaccgggga cggcttcacc ctgcgggagg tgccggtgga cccgcccctg accaaggtgt 3481 tccccgacga cgaatcggac gacgaatcgg acgacgggga ggacggcgac ccggactccc 3541 ggacgttcgt cgcgtacggg gacgacggcg acctggcggg cttcgtggtc gtctcgtact 3601 ccggctggaa ccgccggctg accgtcgagg acatcgaggt cgccccggag caccgggggc 3661 acggggtcgg gcgcgcgttg atggggctcg cgacggagtt cgcccgcgag cggggcgccg 3721 ggcacctctg gctggaggtc accaacgtca acgcaccggc gatccacgcg taccggcgga 3781 tggggttcac cctctgcggc ctggacaccg ccctgtacga cggcaccgcc tcggacggcg 3841 agcaggcgct ctacatgagc atgccctgcc cctaatcagt actgacaata aaaagattct 3901 tgttttcaag aacttgtcat ttgtatagtt tttttatatt gtagttgttc tattttaatc 3961 aaatgttagc gtgatttata ttttttttcg cctcgacatc atctgcccag atgcgaagtt 4021 aagtgcgcag aaagtaatat catgcgtcaa tcgtatgtga atgctggtcg ctatactgct 4081 gtcgattcga tactaacgcc gccatccagt gtcgaaaacg agctcgaatt cctgggtcct 4141 tttcatcacg tgctataaaa ataattataa tttaaatttt ttaatataaa tatataaatt 4201 aaaaatagaa agtaaaaaaa gaaattaaag aaaaaatagt ttttgttttc cgaagatgta 4261 aaagactcta gggggatcgc caacaaatac taccttttat cttgctcttc ctgctctcag 4321 gtattaatgc cgaattgttt catcttgtct gtgtagaaga ccacacacga aaatcctgtg 4381 attttacatt ttacttatcg ttaatcgaat gtatatctat ttaatctgct tttcttgtct 4441 aataaatata tatgtaaagt acgctttttg ttgaaatttt ttaaaccttt gtttattttt 4501 ttttcttcat tccgtaactc ttctaccttc tttatttact ttctaaaatc caaatacaaa 4561 acataaaaat aaataaacac agagtaaatt cccaaattat tccatcatta aaagatacga 4621 ggcgcgtgta agttacaggc aagcgatccg tccgatatca tcagatccac tagtggccta 4681 tgcggccgcg gatctgccgg tctccctata gtgagtcgta ttaatttcga taagccaggt 4741 taacctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc 4801 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc 4861 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa 4921 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt 4981 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg 5041 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg 5101 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag 5161 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc 5221 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa 5281 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg 5341 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc 5401 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac 5461 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg 5521 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt 5581 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt 5641 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa 5701 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga 5761 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt 5821 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg 5881 agacccacgt tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga 5941 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga 6001 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg 6061 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc 6121 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc 6181 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca 6241 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac 6301 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg 6361 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc 6421 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg 6481 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac 6541 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat 6601 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata 6661 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa 6721 agtgccacct gacgtcatct atattaccct gttatcccta gcggatctgc cggtagaggt 6781 gtggtcaata agagcgacct catactatac ctgagaaagc aacctgacct acaggaaaga 6841 gttactcaag aataagaatt ttcgttttaa aacctaagag tcactttaaa atttgtatac 6901 acttattttt tttataactt atttaataat aaaaatcata aatcataaga aattcgctta 6961 tttagaagtg gcgcgccggt ccgttacttg tacagctcgt ccatgccgcc ggtggagtgg 7021 cggccctcgg cgcgttcgta ctgttccacg atggtgtagt cctcgttgtg ggaggtgatg 7081 tccaacttga tgttgacgtt gtaggcgccg ggcagctgca cgggcttctt ggccttgtag 7141 gtggtcttga cctcagcgtc gtagtggccg ccgtccttca gcttcagcct ctgcttgatc 7201 tcgcccttca gggcgccgtc ctcggggtac atccgctcgg aggaggcctc ccagcccatg 7261 gtcttcttct gcattacggg gccgtcggag gggaagttgg tgccgcgcag cttcaccttg 7321 tagatgaact cgccgtcctg cagggaggag tcctgggtca cggtcaccac gccgccgtcc 7381 tcgaagttca tcacgcgctc ccacttgaag ccctcgggga aggacagctt caagtagtcg 7441 gggatgtcgg cggggtgctt cacgtaggcc ttggagccgt acatgaactg aggggacagg 7501 atgtcccagg cgaagggcag ggggccaccc ttggtcacct tcagcttggc ggtctgggtg 7561 ccctcgtagg ggcggccctc gccctcgccc tcgatctcga actcgtggcc gttcacggag 7621 ccctccatgt gcaccttgaa gcgcatgaac tccttgatga tggccatgtt atcctcctcg 7681 cccttgctca ccatggtact agtgtttagt taattatagt tcgttgaccg tatattctaa 7741 aaacaagtac tccttaaaaa aaaaccttga agggaataaa caagtagaat agatagagag 7801 aaaaatagaa aatgcaagag aatttatata ttagaaagag agaaagaaaa atggaaaaaa 7861 aaaaatagga aaagccagaa atagcactag aaggagcgac accagaaaag aaggtgatgg 7921 aaccaattta gctatatata gttaactacc ggctcgatca tctctgcctc cagcatagtc 7981 gaagaagaat tttttttttc ttgaggcttc tgtcagcaac tcgtattttt tctttctttt 8041 ttggtgagcc taaaaagttc ccacgttctc ttgtacgacg ccgtcacaaa caaccttatg 8101 ggtaatttgt cgcggtctgg gtgtataaat gtgtgggtgc aacatgaatg tacggaggta 8161 gtttgctgat tggcggtcta tagatacctt ggttatggcg ccctcacagc cggcaggg //