LOCUS Exported 6959 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION EasyClone system-based yeast integrative vector carrying loxP-flanked KlLEU2 marker, integration into S. cerevisiae XII-2 chromosomal location, USER site for cloning, amp resistance. ACCESSION . VERSION . KEYWORDS pCfB2192 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6959) AUTHORS Stovicek V, Borja GM, Forster J, Borodina I TITLE EasyClone 2.0: expanded toolkit of integrative vectors for stable gene expression in industrial Saccharomyces cerevisiae strains. JOURNAL J Ind Microbiol Biotechnol. 2015 Sep 16. PUBMED 26376869 REFERENCE 2 (bases 1 to 6959) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6959 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 30..219 /gene="S. cerevisiae CYC1" /label=CYC1 terminator /note="transcription terminator for CYC1" protein_bind complement(267..300) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." protein_bind complement(2589..2622) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(2665..2684) /label=T7 /note="T7 promoter, forward primer" promoter complement(2666..2684) /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind complement(2694..2710) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(2703..2725) /label=M13/pUC Forward /note="In lacZ gene" protein_bind complement(3372..3405) /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(3875..3891) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3990..4007) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(4161..4749) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(4241..4260) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(4873..5733) /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 5496..5515 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5734..5825) /gene="bla" /label=AmpR promoter primer_bind 5867..5886 /label=Kan-R /note="Kanamycin resistance gene, reverse primer" primer_bind 5892..5911 /label=pENTR-R /note="pENTR vectors, reverse primer" protein_bind 6118..6151 /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)" terminator 6765..6930 /gene="S. cerevisiae ADH1" /label=ADH1 terminator /note="transcription terminator for the S. cerevisiae alcohol dehydrogenase 1 (ADH1) gene" primer_bind 6851..6875 /label=pHybLex-R /note="ADH terminator, reverse primer" ORIGIN 1 caacggaatg cgtgcgatcg cgtgcattca tccgctctaa ccgaaaagga aggagttaga 61 caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta 121 tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata 181 ctgaaaacct tgcttgagaa ggttttggga cgctcgaaga tcgcgtcagc tgaagcttcg 241 tacgctgcag gtcgacaacc cttaatataa cttcgtataa tgtatgctat acgaagttat 301 taggtctaga gatccgcagg ctaaccggaa cctgtattat ttagtttatg ctacgttaaa 361 taaagacctt tcgttcacat aactgaatgt gtaatggcct tgagatttca agcataccaa 421 gttggtggag acggggtcgt tacaaaagac tctttaagcc aagatttcct tgacagcctt 481 ggcgatagca tcgccaacct cagtggtaga gttagaacca ccaaggtcac cggttctgac 541 acctgcatcc aagacatttc taacagcttc ttcaagagcc ctaccttctt caaccaaatc 601 caaggataac ttcaacatca tagctgcaga taagatggta gcaattgggt taaccttgtt 661 tgctggtaaa tctggggcag aaccatgaca tggttcgtac aaaccgaatg ccttgttagt 721 gtcaggtagg gaagctagag atgcagaagg taataaaccc aaagaacctg gaataacaga 781 ggcttcatcg gagataatat caccaaacat gttgttggta ataacaacac cgtttagctt 841 agttggtgat ttaaccaaaa tcatagcagc agagtcgatc aattggtgct gaacagttaa 901 ttgtgggaac tcagtcttga tggtttcttc aacagtcttt ctccacaatc tggaagaggc 961 aagcacgtta gccttgtcaa gtgaccagat tggtaatggt gggttttgtt gcaatgccaa 1021 gaaagcagcc attcttgtaa ttctttgaac ttcaggaaca ctgtatttct cagagtccca 1081 agcaactccg tcaccttcat cttcttttct ttcaccaaag tagataccac caaccaattc 1141 tctaacgacg acgaaatcgg taccctttgc atattcaggc ttcaaaggag aaagatctag 1201 taaagaatca gaagcaaagt tacatggtct taagttggcg tatagaccca attcctttct 1261 gatcttcaat agaccttgtt ctggtctaac tgcgcccgta ccccattttg gaccaccaac 1321 agcacctagt aagacagcat cggctttctt agaggcttct agagcttcat ctggtaaagg 1381 agtgccagtg gcatcgatgg cagcaccccc gatcaagtga tgttggaaat tgaacttaat 1441 ttctggacgg acttcagcaa tggcattcaa gaccttaata gcttcgtcag taacttcttt 1501 accgacgtga tcacccggta ggacaacgat attcttagac atggtataat ctgtgtagtg 1561 tgggatactt tttacttctt caaataggta tcaacaataa aaataaatcg aagcaaatgt 1621 aggaatgcgt taaagcagat gtacttactt agagtacata gatatattta tataattcaa 1681 tatataaaag tatatgaagc atctgatgtt gaacctgtca tgactctaaa ctggaatggg 1741 tagttatgct tctgaaggtt ttccgttaga aaatgagtgc agggatcagg cccggaaccg 1801 gttttagcct gaaaggaaaa gaaaaaaatg cggtgattac tagcacgtga ctgcgctgaa 1861 ttggaatcca ccacaagtac agcgaatggc aagagggaag gggaatatat tgtgcctggc 1921 ggtagtctgc tatatcaaat gttaaacata actaaatgca gtagtgaata gagtaatgta 1981 tttctacata tgtgggtagg gccaatggga gcccgatgtc tattgtacag atattttcct 2041 tttatagtta atcaattcaa cttggaaatc attgagtcga tatctacttg gataactgga 2101 gccgtggcag cgtttgaaat ttgacgagtc tccttgtttt gttcgcgcat tttcagttta 2161 ttcaaacggg agatggcacc gaatccactg acttctctgt tcaatttgaa tggtaatagg 2221 aagaaagtac atttcgagaa attttcgccc aacgtcgaaa tctcatggat gatatcctcg 2281 atacaaatga caccttcatc acctaatttc tcttcaacga tattgttgtc attcaagatt 2341 atctcttttg gttctttatc ttctggcctt tgccacatta ttctagatct cttttggatt 2401 aaagaacgga tagatgcaag tgaaggttgt ccgataacaa tatatggtga tacaagtttc 2461 aacaatgggt acacgtcctt ggtaagtttg atgaaaacac cagtgttcaa ctcaacaagt 2521 ctcaaaagag ataacacttt gtaagccttt gctgggatct tcacagcgag ctctcgagaa 2581 cccttaatat aacttcgtat aatgtatgct atacgaagtt attaggtgat atcagatcca 2641 ctagtggcct atgcacccaa ttcgccctat agtgagtcgt attacgcgcg ctcactggcc 2701 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccctgcagg actagtgctg 2761 aggcattaat ctactatcgg cgactctctc gaaatttttc ttaacgcgtc cttgtactgc 2821 gtctaacgct tttgccactt ggatttctat tataggaaat agtctcactt actgggcgac 2881 gaattttcgc gttttgatga agcacaggaa gaatttcttt tttttttggc ttcttctggt 2941 tccgtttttt acgcgcacaa atctaaaaaa agaaataatt ataacctagt ctcgaaaatt 3001 ttcatcgatc cattcgttcc tttttttcga ttttttcaga tcaaaattct tgtttctttc 3061 tttgtcttag tttatattaa aagatatttt gattttactc ctgaactatt tattctttct 3121 aagaaggcca gaacactaca gctgttttaa ccgactacga agttctccat tctcgaacac 3181 tagccttcat ttaccaaaca ggaactagcg tatatcatta gtccttattc gaaaagagat 3241 tggtagatat ttattgtagt ttgtgagaag gagaaaatac tgtcattgga ctgatagtta 3301 gaggacatta acctctctta cgttcgctca ttaagacctc agcgcggccg caaatttaaa 3361 taaaatgaag tgaagttcct atactttcta gagaatagga acttctatag tgagtcgaat 3421 aagggcgaca caaaatttat tctaaatgca taataaatac tgataacatc ttatagtttg 3481 tattatattt tgtattatcg ttgacatgta taattttgat atcaaaaact gattttccct 3541 ttattatttt cgagatttat tttcttaatt ctctttaaca aactagaaat attgtatata 3601 caaaaaatca taaataatag atgaatagtt taattatagg tgttcatcaa tcgaaaaagc 3661 aacgtatctt atttaaagtg cgttgctttt ttctcattta taaggttaaa taattctcat 3721 atatcaagca aagtgacagg cgcccttaaa tattctgaca aatgctcttt ccctaaactc 3781 cccccataaa aaaacccgcc gaagcgggtt tttacgttat ttgcggatta acgattactc 3841 gttatcagaa ccgcccaggg ggcccgagct taagactggc cgtcgtttta caacacagaa 3901 agagtttgta gaaacgcaaa aaggccatcc gtcaggggcc ttctgcttag tttgatgcct 3961 ggcagttccc tactctcgcc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt 4021 cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca 4081 ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa 4141 aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat 4201 cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc 4261 cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc 4321 gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt 4381 tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac 4441 cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg 4501 ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca 4561 gagttcttga agtggtgggc taactacggc tacactagaa gaacagtatt tggtatctgc 4621 gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa 4681 accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa 4741 ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg gaacgacgcg 4801 cgcgtaactc acgttaaggg attttggtca tgagcttgcg ccgtcccgtc aagtcagcgt 4861 aatgctctgc ttttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt 4921 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt 4981 accatctggc cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt 5041 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc 5101 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa 5161 tagtttgcgc aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg 5221 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt 5281 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc 5341 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt 5401 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg 5461 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac 5521 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc 5581 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt 5641 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg 5701 aataagggcg acacggaaat gttgaatact catattcttc ctttttcaat attattgaag 5761 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa 5821 acaaataggg gtcagtgtta caaccaatta accaattctg aacattatcg cgagcccatt 5881 tatacctgaa tatggctcat aacacccctt gtttgcctgg cggcagtagc gcggtggtcc 5941 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtgggga 6001 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa 6061 gactgggcct ttcgcccggg ctaattatgg ggtgtcgccc ttattcgact ctatagtgaa 6121 gttcctattc tctagaaagt ataggaactt ctgaagtggg gatttaaatg cggccgcgct 6181 gagggtttaa tcggcatgca aacatctaca caattagcaa gggcaatcca tattttgtct 6241 tttcgcgccc tggaaaggcc taagtaatgt cgtaaacgca ttctatctgt acttcaactc 6301 tcctctgtgc attggtttgt gcaaatcaca ttttacgata ctgccagata tatgcaaaaa 6361 gagaaaacca agggaccaga acaaagcaaa attacgatat tcttcgaatt ccttcgtgct 6421 tgactaagac aaagggatgg acgtagcgat ttttagcggg ccaagaactg gttccgaaaa 6481 agcacaggta caccgaaccc tcagctaagg agggacagca ccgatgcgga aggacaaact 6541 ttctttttgc ctatcacagt atcttatcga gctaactatt ttcgacacac atgaaaaagc 6601 agaaatatta acgaaaaaga aaagaaagac catgtcatgt acgggcaatc agaatctgta 6661 acaagcgcca tttttttttc tgtatcgggc cctccttact gctctccttc cgtgtaacgc 6721 gttatgatta agtcctcagc gagctcgcat ggaatgcgtg cgatgagcga cctcatgcta 6781 tacctgagaa agcaacctga cctacaggaa agagttactc aagaataaga attttcgttt 6841 taaaacctaa gagtcacttt aaaatttgta tacacttatt ttttttataa cttatttaat 6901 aataaaaatc ataaatcata agaaattcgc ttatttagaa gtgtcaacaa cgtatctac //