LOCUS Exported 6932 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION (parent plasmid pCFB262) pXII - 4 -LoxP -SpHIS5 -209bp - pCYC1 -PcaO -108 -yeGFP. ACCESSION . VERSION . KEYWORDS pFA197 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6932) AUTHORS Ambri F, D'Ambrosio V, Di Blasi R, Maury J, Jacobsen SAB, McCloskey D, Jensen MK, Keasling JD TITLE High-Resolution Scanning of Optimal Biosensor Reporter Promoters in Yeast. JOURNAL ACS Synth Biol. 2020 Feb 21;9(2):218-226. doi: 10.1021/acssynbio.9b00333. Epub 2020 Jan 31. PUBMED 31935067 REFERENCE 2 (bases 1 to 6932) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6932 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 178..343 /gene="S. cerevisiae ADH1" /label=ADH1 terminator /note="transcription terminator for the S. cerevisiae alcohol dehydrogenase 1 (ADH1) gene" primer_bind 264..288 /label=pHybLex-R /note="ADH terminator, reverse primer" CDS 695..1411 /codon_start=1 /product="yeast-enhanced green fluorescent protein" /label=yeGFP /note="codons optimized for translation in S. cerevisiae and C. albicans" /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" terminator 1425..1614 /gene="S. cerevisiae CYC1" /label=CYC1 terminator /note="transcription terminator for CYC1" protein_bind complement(1662..1695) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." gene 1750..2950 /label=HIS3MX6 /note="yeast selectable marker encoding the S. pombe his5 gene, which corresponds to S. cerevisiae HIS3" promoter 1750..2093 /label=TEF promoter /note="Ashbya gossypii TEF promoter" CDS 2094..2747 /codon_start=1 /gene="Schizosaccharomyces pombe his5" /product="imidazoleglycerol-phosphate dehydratase, required for histidine biosynthesis" /label=S. pombe his5 /note="yeast auxotrophic marker; corresponds to S. cerevisiae HIS3" /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQ KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ AMGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL" terminator 2753..2950 /label=TEF terminator /note="Ashbya gossypii TEF terminator" protein_bind complement(3010..3043) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(3086..3105) /label=T7 /note="T7 promoter, forward primer" promoter complement(3087..3105) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(3115..3131) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3124..3146) /label=M13/pUC Forward /note="In lacZ gene" protein_bind complement(3751..3784) /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" primer_bind complement(4254..4270) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4369..4386) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4540..5128) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4620..4639) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(5250..6110) /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 5873..5892 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6111..6202) /gene="bla" /label=AmpR promoter primer_bind 6244..6263 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 6269..6288 /label=pENTR-R /note="pENTR vectors, reverse primer" protein_bind 6495..6528 /label=FRT (minimal) /bound_moiety="FLP recombinase from the Saccharomyces cerevisiae 2u plasmid" /note="supports FLP-mediated excision but not integration (Turan and Bode, 2011)" ORIGIN 1 tggatttgtt ttatcaatac acagtctaca gtcaagaatt ttttttatca aattttgcgt 61 tcgagcgtat aaaatagccg ctgtagctac ttaagttcct gttcagcgat agtttttttc 121 catcacacgt actatggcaa ttaagtcctc agcgagctcg catggaatgc gtgcgatgag 181 cgacctcatg ctatacctga gaaagcaacc tgacctacag gaaagagtta ctcaagaata 241 agaattttcg ttttaaaacc taagagtcac tttaaaattt gtatacactt atttttttta 301 taacttattt aataataaaa atcataaatc ataagaaatt cgcttattta gaagtgtcaa 361 caacgtatct accaacggaa tacgtgcgat ccaggcaact ttagtgctga cacatacagg 421 catatatata tgtgtgcgac gacacatgat catatggcat gcatgtgctc tgtatgtata 481 taaaactctt ggatcgtata acctcctggt taagggaaag ccacgaaata tcattttacc 541 taaccggatg aaacatccaa atctgacgac gttttcttct tttctctaaa tattctttcc 601 ttatacatta ggacctttgc agcataaatt actatacttc tatagacaca caaacacaaa 661 tacacacact aaattaataa tctgtcataa aacaatgtct aaaggtgaag aattattcac 721 tggtgttgtc ccaattttgg ttgaattaga tggtgatgtt aatggtcaca aattttctgt 781 ctccggtgaa ggtgaaggtg atgctactta cggtaaattg accttaaaat ttatttgtac 841 tactggtaaa ttgccagttc catggccaac cttagtcact actttcggtt atggtgttca 901 atgttttgcg agatacccag atcatatgaa acaacatgac tttttcaagt ctgccatgcc 961 agaaggttat gttcaagaaa gaactatttt tttcaaagat gacggtaact acaagaccag 1021 agctgaagtc aagtttgaag gtgatacctt agttaataga atcgaattaa aaggtattga 1081 ttttaaagaa gatggtaaca ttttaggtca caaattggaa tacaactata actctcacaa 1141 tgtttacatc atggctgaca aacaaaagaa tggtatcaaa gttaacttca aaattagaca 1201 caacattgaa gatggttctg ttcaattagc tgaccattat caacaaaata ctccaattgg 1261 tgatggtcca gtcttgttac cagacaacca ttacttatcc actcaatctg ccttatccaa 1321 agatccaaac gaaaagagag accacatggt cttgttagaa tttgttactg ctgctggtat 1381 tacccatggt atggatgaat tgtacaaata aatcgcgtgc attcatccgc tctaaccgaa 1441 aaggaaggag ttagacaacc tgaagtctag gtccctattt atttttttat agttatgtta 1501 gtattaagaa cgttatttat atttcaaatt tttctttttt ttctgtacag acgcgtgtac 1561 gcatgtaaca ttatactgaa aaccttgctt gagaaggttt tgggacgctc gaagatcgcg 1621 tcagctgaag cttcgtacgc tgcaggtcga caacccttaa tataacttcg tataatgtat 1681 gctatacgaa gttattaggt ctagagatct gtttagcttg cctcgtcccc gccgggtcac 1741 ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt gcgcagctca 1801 ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt ataatcattt 1861 gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc ctcgctgcag 1921 acctgcgagc agggaaacgc tcccctcaca gacgcgttga attgtcccca cgccgcgccc 1981 ctgtagagaa atataaaagg ttaggatttg ccactgaggt tcttctttca tatacttcct 2041 tttaaaatct tgctaggata cagttctcac atcacatccg aacataaaca accatgggta 2101 ggagggcttt tgtagaaaga aatacgaacg aaacgaaaat cagcgttgcc atcgctttgg 2161 acaaagctcc cttacctgaa gagtcgaatt ttattgatga acttataact tccaagcatg 2221 caaaccaaaa gggagaacaa gtaatccaag tagacacggg aattggattc ctggatcaca 2281 tgtatcatgc actggctaaa catgcaggct ggagcttacg actttactca agaggtgatt 2341 taatcatcga tgatcatcac actgcagaag atactgctat tgcacttggt attgcattca 2401 agcaggctat gggtaacttt gccggcgtta aaagatttgg acatgcttat tgtccacttg 2461 acgaagccct ttctagaagc gtagttgact tgtcgggacg gccctatgct gttatcgatt 2521 tgggattaaa gcgtgaaaag gttggggaat tgtcctgtga aatgatccct cacttactat 2581 attccttttc ggtagcagct ggaattactt tgcatgttac ctgcttatat ggtagtaatg 2641 accatcatcg tgctgaaagc gcttttaaat ctctggctgt tgccatgcgc gcggctacta 2701 gtcttactgg aagttctgaa gtcccaagca cgaagggagt gttgtaaaga gtactgacaa 2761 taaaaagatt cttgttttca agaacttgtc atttgtatag tttttttata ttgtagttgt 2821 tctattttaa tcaaatgtta gcgtgattta tatttttttt cgcctcgaca tcatctgccc 2881 agatgcgaag ttaagtgcgc agaaagtaat atcatgcgtc aatcgtatgt gaatgctggt 2941 cgctatactg ctgtcgattc gatactaacg ccgccatcca gtttaaacga gctctcgaga 3001 acccttaata taacttcgta taatgtatgc tatacgaagt tattaggtga tatcagatcc 3061 actagtggcc tatgcaccca attcgcccta tagtgagtcg tattacgcgc gctcactggc 3121 cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccctgcag gactagtgct 3181 gaggcattaa tccaactttg tactattcct tccccgttta ctcaattctt gaagccaatt 3241 tgtacaattc cccattagag tcaaataaaa ggatgcctca cggaggtatg ttacccgcgc 3301 tatttcacat ggctcattga attagaggtg gaatttggtg taccctcccc tcctcatctg 3361 atgaagtagt gatccgacaa ttcttaaaag ttgtagacat tacttttacc accaactaag 3421 ttgtatttat attgctaccc ttatcctttt atatctaact agcgctcata aggttggggc 3481 aatactaaaa ctgtgttctt attcaactca ttaaatacgt ggcagtacgt accctattag 3541 aaacaatagg aaacagcaga gtcggaagaa gccaaatgcc agatttgaag tccaaaacct 3601 tgtcaagcca atctttggga gcggctattc ctccagaaat tgtgtaccaa atacttacat 3661 accagtttag ggatttgtta agaaatgacc atccaggtac ggcagaaaat taagacctca 3721 gcgcggccgc aaatttaaat aaaatgaagt gaagttccta tactttctag agaataggaa 3781 cttctatagt gagtcgaata agggcgacac aaaatttatt ctaaatgcat aataaatact 3841 gataacatct tatagtttgt attatatttt gtattatcgt tgacatgtat aattttgata 3901 tcaaaaactg attttccctt tattattttc gagatttatt ttcttaattc tctttaacaa 3961 actagaaata ttgtatatac aaaaaatcat aaataataga tgaatagttt aattataggt 4021 gttcatcaat cgaaaaagca acgtatctta tttaaagtgc gttgcttttt tctcatttat 4081 aaggttaaat aattctcata tatcaagcaa agtgacaggc gcccttaaat attctgacaa 4141 atgctctttc cctaaactcc ccccataaaa aaacccgccg aagcgggttt ttacgttatt 4201 tgcggattaa cgattactcg ttatcagaac cgcccagggg gcccgagctt aagactggcc 4261 gtcgttttac aacacagaaa gagtttgtag aaacgcaaaa aggccatccg tcaggggcct 4321 tctgcttagt ttgatgcctg gcagttccct actctcgcct tccgcttcct cgctcactga 4381 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 4441 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 4501 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 4561 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 4621 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 4681 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 4741 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 4801 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 4861 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 4921 gtatgtaggc ggtgctacag agttcttgaa gtggtgggct aactacggct acactagaag 4981 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 5041 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 5101 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 5161 cgctcagtgg aacgacgcgc gcgtaactca cgttaaggga ttttggtcat gagcttgcgc 5221 cgtcccgtca agtcagcgta atgctctgct taccaatgct taatcagtga ggcacctatc 5281 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact 5341 acgatacggg agggcttacc atctggcccc agcgctgcga tgataccgcg agaaccacgc 5401 tcaccggctc cggatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt 5461 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta 5521 agtagttcgc cagttaatag tttgcgcaac gttgttgcca tcgctacagg catcgtggtg 5581 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt 5641 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc 5701 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt 5761 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc 5821 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc 5881 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa 5941 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac 6001 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa 6061 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat attcttcctt 6121 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa 6181 tgtatttaga aaaataaaca aataggggtc agtgttacaa ccaattaacc aattctgaac 6241 attatcgcga gcccatttat acctgaatat ggctcataac accccttgtt tgcctggcgg 6301 cagtagcgcg gtggtcccac ctgaccccat gccgaactca gaagtgaaac gccgtagcgc 6361 cgatggtagt gtggggactc cccatgcgag agtagggaac tgccaggcat caaataaaac 6421 gaaaggctca gtcgaaagac tgggcctttc gcccgggcta attatggggt gtcgccctta 6481 ttcgactcta tagtgaagtt cctattctct agaaagtata ggaacttctg aagtggggat 6541 ttaaatgcgg ccgcgctgag ggtttaatgt atccggctgt tccttcatag ccctttcaat 6601 gaacgttgca gccctttgaa gattggccat tttgtcagga ctcgagcctg acagttggac 6661 caacgcaact ttaatttttt gtgaaagaat cttcgaagca ctcatactgg cgatcttcac 6721 gccctcctgc tattacaaaa gctgtgtttt tacaagaatc aaattaagtt agcaagatat 6781 tatacaacat tattgataat ttcaatatcg tgttcgtacc tgatgacgta tctgtgcatt 6841 gataaggccc gcatggtttc agaaagcaga gcggaacgat tccaaattag tggccttgtg 6901 ctttgcatgt caattgtgtt accttcagct cg //