LOCUS Exported 6267 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION synthetic circular DNA ACCESSION . VERSION . KEYWORDS pDZ264 (pKAN tdtomato cyc loxP KAN loxP) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6267) AUTHORS Larson DR, Zenklusen D, Wu B, Chao JA, Singer RH TITLE Real-time observation of transcription initiation and elongation on an endogenous yeast gene. JOURNAL Science. 2011 Apr 22;332(6028):475-8. PUBMED 21512033 REFERENCE 2 (bases 1 to 6267) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6267 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(20..36) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(21..37) /label=pBluescriptKS /note="For pBluescript vector" terminator 39..286 /gene="S. cerevisiae CYC1" /label=CYC1 terminator /note="transcription terminator for CYC1" primer_bind complement(55..73) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" protein_bind complement(337..370) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." gene 425..1781 /label=kanMX /note="yeast selectable marker conferring kanamycin resistance (Wach et al., 1994)" promoter 425..768 /label=TEF promoter /note="Ashbya gossypii TEF promoter" CDS 769..1578 /codon_start=1 /gene="aph(3')-Ia" /product="aminoglycoside phosphotransferase" /label=KanR /note="confers resistance to kanamycin" /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" primer_bind complement(836..855) /label=Kan-R /note="Kanamycin resistance gene, reverse primer" terminator 1584..1781 /label=TEF terminator /note="Ashbya gossypii TEF terminator" protein_bind complement(1844..1877) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." primer_bind complement(1928..1948) /label=T3 /note="T3 promoter, forward primer" promoter complement(1928..1946) /label=T3 promoter /note="promoter for bacteriophage T3 RNA polymerase" primer_bind complement(1967..1983) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(1967..1983) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(1980..2002) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 1991..2007 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /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(2015..2045) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 2060..2081 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(2198..2215) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(2369..2957) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2449..2468) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(3128..3988) /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 3751..3770 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(3989..4093) /gene="bla" /label=AmpR promoter rep_origin 4120..4575 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4207..4226) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4417..4438 /label=F1ori-F /note="F1 origin, forward primer" primer_bind 4701..4723 /label=M13/pUC Forward /note="In lacZ gene" primer_bind 4715..4732 /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind 4716..4732 /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind 4742..4761 /label=T7 /note="T7 promoter, forward primer" promoter 4742..4760 /label=T7 promoter /note="promoter for bacteriophage T7 RNA polymerase" primer_bind 4793..4809 /label=SK primer /note="common sequencing primer, one of multiple similar variants" primer_bind 4793..4809 /label=pBluescriptSK /note="For pBluescript vector" regulatory 4815..4824 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 4821..6248 /codon_start=1 /product="tandem dimeric (pseudo-monomeric) derivative of DsRed (Shaner et al., 2004)" /label=tdTomato /note="mammalian codon-optimized" /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV LKGEIHQALKLKDGGRYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE RSEGRHHLFLYGMDELYK" ORIGIN 1 aattcgatat caagcttatc gataccgtcg acctcgagtc atgtaattag ttatgtcacg 61 cttacattca cgccctcccc ccacatccgc tctaaccgaa aaggaaggag ttagacaacc 121 tgaagtctag gtccctattt atttttttat agttatgtta gtattaagaa cgttatttat 181 atttcaaatt tttctttttt ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa 241 aaccttgctt gagaaggttt tgggacgctc gaaggcttta atttgcggcc ggtacccagc 301 tgaagcttcg tacgctgcag gtcgacaacc cttaatataa cttcgtataa tgtatgctat 361 acgaagttat taggtctaga gatctgttta gcttgcctcg tccccgccgg gtcacccggc 421 cagcgacatg gaggcccaga ataccctcct tgacagtctt gacgtgcgca gctcaggggc 481 atgatgtgac tgtcgcccgt acatttagcc catacatccc catgtataat catttgcatc 541 catacatttt gatggccgca cggcgcgaag caaaaattac ggctcctcgc tgcagacctg 601 cgagcaggga aacgctcccc tcacagacgc gttgaattgt ccccacgccg cgcccctgta 661 gagaaatata aaaggttagg atttgccact gaggttcttc tttcatatac ttccttttaa 721 aatcttgcta ggatacagtt ctcacatcac atccgaacat aaacaaccat gggtaaggaa 781 aagactcacg tttcgaggcc gcgattaaat tccaacatgg atgctgattt atatgggtat 841 aaatgggctc gcgataatgt cgggcaatca ggtgcgacaa tctatcgatt gtatgggaag 901 cccgatgcgc cagagttgtt tctgaaacat ggcaaaggta gcgttgccaa tgatgttaca 961 gatgagatgg tcagactaaa ctggctgacg gaatttatgc ctcttccgac catcaagcat 1021 tttatccgta ctcctgatga tgcatggtta ctcaccactg cgatccccgg caaaacagca 1081 ttccaggtat tagaagaata tcctgattca ggtgaaaata ttgttgatgc gctggcagtg 1141 ttcctgcgcc ggttgcattc gattcctgtt tgtaattgtc cttttaacag cgatcgcgta 1201 tttcgtctcg ctcaggcgca atcacgaatg aataacggtt tggttgatgc gagtgatttt 1261 gatgacgagc gtaatggctg gcctgttgaa caagtctgga aagaaatgca taagcttttg 1321 ccattctcac cggattcagt cgtcactcat ggtgatttct cacttgataa ccttattttt 1381 gacgagggga aattaatagg ttgtattgat gttggacgag tcggaatcgc agaccgatac 1441 caggatcttg ccatcctatg gaactgcctc ggtgagtttt ctccttcatt acagaaacgg 1501 ctttttcaaa aatatggtat tgataatcct gatatgaata aattgcagtt tcatttgatg 1561 ctcgatgagt ttttctaatc agtactgaca ataaaaagat tcttgttttc aagaacttgt 1621 catttgtata gtttttttat attgtagttg ttctatttta atcaaatgtt agcgtgattt 1681 atattttttt tcgcctcgac atcatctgcc cagatgcgaa gttaagtgcg cagaaagtaa 1741 tatcatgcgt caatcgtatg tgaatgctgg tcgctatact gctgtcgatt cgatactaac 1801 gccgccatcc agtgtcgaaa acgagctctc gagaaccctt aatataactt cgtataatgt 1861 atgctatacg aagttattag gtgatatcag atccactagt ggcctatgcg gtacccagct 1921 tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc 1981 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt 2041 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc 2101 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg 2161 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct 2221 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca 2281 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga 2341 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc 2401 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg 2461 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat 2521 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt 2581 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc 2641 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg 2701 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg 2761 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg 2821 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg 2881 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca 2941 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga 3001 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga 3061 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt 3121 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt 3181 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat 3241 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag 3301 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct 3361 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt 3421 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg 3481 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca 3541 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt 3601 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat 3661 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac 3721 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa 3781 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt 3841 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt 3901 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa 3961 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt 4021 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa 4081 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgcgccctgt agcggcgcat 4141 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag 4201 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc 4261 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc 4321 ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga tagacggttt 4381 ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa 4441 caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg ccgatttcgg 4501 cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat 4561 taacgcttac aatttccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt 4621 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag 4681 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgagcgcgc 4741 gtaatacgac tcactatagg gcgaattgga gctccaccgc ggtggcggcc gctctagaac 4801 tagtggatcc ggtcgccacc atggtgagca agggcgagga ggtcatcaaa gagttcatgc 4861 gcttcaaggt gcgcatggag ggctccatga acggccacga gttcgagatc gagggcgagg 4921 gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc aagggcggcc 4981 ccctgccctt cgcctgggac atcctgtccc cccagttcat gtacggctcc aaggcgtacg 5041 tgaagcaccc cgccgacatc cccgattaca agaagctgtc cttccccgag ggcttcaagt 5101 gggagcgcgt gatgaacttc gaggacggcg gtctggtgac cgtgacccag gactcctccc 5161 tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg caccaacttc ccccccgacg 5221 gccccgtaat gcagaagaag accatgggct gggaggcctc caccgagcgc ctgtaccccc 5281 gcgacggcgt gctgaagggc gagatccacc aggccctgaa gctgaaggac ggcggccact 5341 acctggtgga gttcaagacc atctacatgg ccaagaagcc cgtgcaactg cccggctact 5401 actacgtgga caccaagctg gacatcacct cccacaacga ggactacacc atcgtggaac 5461 agtacgagcg ctccgagggc cgccaccacc tgttcctggg gcatggcacc ggcagcaccg 5521 gcagcggcag ttccggcacc gcttcttccg aggacaacaa catggccgtc atcaaagagt 5581 tcatgcgctt caaggtgcgc atggagggct ccatgaacgg ccacgagttc gagatcgagg 5641 gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg 5701 gcggccccct gcccttcgcc tgggacatcc tgtcccccca gttcatgtac ggctccaagg 5761 cgtacgtgaa gcaccccgcc gacatccccg attacaagaa gctgtccttc cccgagggct 5821 tcaagtggga gcgcgtgatg aacttcgagg acggcggtct ggtgaccgtg acccaggact 5881 cctccctgca ggacggcacg ctgatctaca aggtgaagat gcgcggcacc aacttccccc 5941 ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctccacc gagcgcctgt 6001 acccccgcga cggcgtgctg aagggcgaga tccaccaggc cctgaagctg aaggacggcg 6061 gccgctacct ggtggagttc aagaccatct acatggccaa gaagcccgtg caactgcccg 6121 gctactacta cgtggacacc aagctggaca tcacctccca caacgaggac tacaccatcg 6181 tggaacagta cgagcgctcc gagggccgcc accacctgtt cctgtacggc atggacgagc 6241 tgtacaagtc cggaatatat aagggcg //