LOCUS Exported 6562 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Yeast Expression plasmid for SARS-CoV-2 helicase. ACCESSION . VERSION . KEYWORDS pGBW-m4046232 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6562) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6562) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6562 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 10..417 /gene="S. cerevisiae TEF1" /label=TEF1 promoter /note="promoter for EF-1-alpha" CDS 2237..2254 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 2255..2272 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(2317..2335) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 2686..3489 /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" rep_origin 3572..4554 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 4632..4661 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 4683..5270 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 5282..5309 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(5333..5524) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(5525..6385) /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESLRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLATIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGSLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind 6148..6167 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6386..6488) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 6489..6520 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" ORIGIN 1 gcggccgccc cacacaccat agcttcaaaa tgtttctact ccttttttac tcttccagat 61 tttctcggac tccgcgcatc gccgtaccac ttcaaaacac ccaagcacag catactaaat 121 ttcccctctt tcttcctcta gggtgtcgtt aattacccgt actaaaggtt tggaaaagaa 181 aaaagagacc gcctcgtttc tttttcttcg tcgaaaaagg caataaaaat ttttatcacg 241 tttctttttc ttgaaaattt ttttttttga tttttttctc tttcgatgac ctcccattga 301 tatttaagtt aataaacggt cttcaatttc tcaagtttca gtttcatttt tcttgttcta 361 ttacaacttt ttttacttct tgctcattag aaagaaagca tagcaatcta atctaagttt 421 taattacaaa atggcagtcg gcgcttgtgt attatgcaac tcccagacgt cgttgagatg 481 tggtgcttgt attcgtagac ctttcctttg ctgtaaatgt tgctatgacc acgttatctc 541 aacttctcat aagctagttc tgagcgtcaa tccatacgtg tgtaacgccc ccgggtgtga 601 tgttacagac gttacccaat tgtacttggg aggtatgtct tactattgta agtcccacaa 661 gccacctata tcttttccat tatgcgctaa cggtcaagtc ttcggtttgt acaaaaatac 721 ctgtgttggt agtgataatg ttaccgactt taacgccatt gctacttgtg attggactaa 781 cgcgggtgat tatatcttgg ctaacacttg taccgagagg cttaagttgt tcgccgctga 841 aaccttaaag gctacggaag aaacattcaa attgagttac ggtatcgcta ctgtcagaga 901 agttctatct gatagagaat tgcatctctc ctgggaggtg ggtaagccaa gaccaccgtt 961 gaaccggaat tacgtcttta ctggatacag ggttactaag aactctaagg ttcaaattgg 1021 ggaatatacc tttgaaaagg gcgactacgg tgacgccgta gtctatagag gtaccaccac 1081 ttacaaattg aacgtcggtg actatttcgt tttaaccagc cacacagtga tgccactgtc 1141 agcgccaaca ttagttccgc aagagcatta cgttagaatt actggtttgt atccaactct 1201 aaatatctct gatgaatttt cctccaatgt ggctaactac caaaaagtag gtatgcaaaa 1261 gtactctacc ttgcaaggtc cacctggtac tgggaagtcg cacttcgcta taggtttggc 1321 actgtactac ccatctgcta gaattgttta cacggcatgt tctcatgctg ctgtagacgc 1381 cttgtgtgaa aaggccttga aatatttacc cattgataaa tgttcaagaa ttatcccagc 1441 acgagctcgt gttgaatgtt tcgacaagtt taaggtcaac agcacactag aacaatacgt 1501 cttctgcact gttaatgctt tgccagaaac gactgctgat atcgtcgtct ttgatgaaat 1561 ctcgatggcc actaactatg acttatccgt agttaatgct agacttcgtg caaagcacta 1621 cgtctatatt ggtgatccag ctcagctgcc tgcaccaaga actttgttaa ccaagggtac 1681 cttggaacca gaatacttca actctgtgtg tcgcctgatg aagacaatcg gccctgatat 1741 gttcctaggt acatgtagac gttgtccagc tgaaattgtg gacacggtca gtgccttggt 1801 ttacgataac aaattaaagg cgcataagga caaaagcgct caatgtttca agatgtttta 1861 caagggtgtc attactcacg atgttagttc ggccatcaat agaccccaaa taggtgttgt 1921 tagagaattt ttgacaagga acccagcatg gagaaaggct gttttcatat ccccatacaa 1981 ttctcaaaac gcggtagcct caaaaatctt gggtcttccc actcagaccg ttgattcttc 2041 ccaaggaagt gaatatgact acgtgatttt cactcaaacc accgagactg cccactcttg 2101 caacgtcaat cgtttcaacg ttgctattac gagagccaag gttggcatct tgtgtatcat 2161 gtctgataga gacttgtacg ataaattaca gtttacatcc ttagaaatac caagacgaaa 2221 cgtcgctact ttgcaactcg tgccgagagg ttctcaccat catcaccatc attaaacagg 2281 ccccttttcc tttgtcgata tcatgtaatt agttatgtca cgcttacatt cacgccctcc 2341 tcccacatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat 2401 ttattttttt taatagttat gttagtatta agaacgttat ttatatttca aatttttctt 2461 ttttttctgt acaaagcggc cgccgaacca ctagtgcact gcagtacacg gtttccttga 2521 aatttttttg attcggtaat ctccgaacag aaggaagaac gaaggaagga gcacagactt 2581 agattggtat atatacgcat atgtagtgtt gaagaaacat gaaattgccc agtattctta 2641 acccaactgc acagaacaaa aacgtgcagg aaacgaagat aaatcatgtc gaaagctaca 2701 tataaggaac gtgctgctac tcatcctagt cctgttgctg ccaagctatt taatatcatg 2761 cacgaaaagc aaacaaactt gtgtgcttca ttggatgttc gtaccaccaa ggaattactg 2821 gagttagttg aagcattagg tcccaaaatt tgtttactaa aaacacatgt ggatatcttg 2881 actgattttt ccatggaggg cacagttaag ccgctaaagg cattatccgc caagtacaat 2941 tttttactct tcgaggacag aaaatttgct gacattggta atacagtcaa attgcagtac 3001 tctgcgggtg tatacagaat agcagaatgg gcagacatta cgaatgcaca cggtgtggtg 3061 ggcccaggta ttgttagcgg tttgaagcag gcggcagaag aagtaacaaa ggaacctaga 3121 ggccttttga tgttagcaga attgtcatgc aagggctccc tatctactgg agaatatact 3181 aagggtactg ttgacattgc gaagagcgac aaagattttg ttatcggctt tattgctcaa 3241 agagacatgg gtggaagaga tgaaggttac gattggttga ttatgacacc cggtgtgggt 3301 ttagatgaca agggagatgc attgggtcaa cagtatagaa ccgtggatga tgtggtttct 3361 acaggatctg acattattat tgttggaaga ggactatttg caaagggaag ggatgctaag 3421 gtagagggtg aacgttacag aaaagcaggc tgggaagcat atttgagaag atgcggccag 3481 caaaactaaa aaactgtatt ataagtaaat gcatgtatac taaactcaca aattagagct 3541 tcaatttaat tatatcagtt attacccgag taacgaagca tctgtgcttc attttgtaga 3601 acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag aatctgagct gcatttttac 3661 agaacagaaa tgcaacgcga aagcgctatt ttaccaacga agaatctgtg cttcattttt 3721 gtaaaacaaa aatgcaacgc gagagcgcta atttttcaaa caaagaatct gagctgcatt 3781 tttacagaac agaaatgcaa cgcgagagcg ctattttacc aacaaagaat ctatacttct 3841 tttttgttct acaaaaatgc atcccgagag cgctattttt ctaacaaagc atcttagatt 3901 actttttttc tcctttgtgc gctctataat gcagtctctt gataactttt tgcactgtag 3961 gtccgttaag gttagaagaa ggctactttg gtgtctattt tctcttccat aaaaaaagcc 4021 tgactccact tcccgcgttt actgattact agcgaagctg cgggtgcatt ttttcaagat 4081 aaaggcatcc ccgattatat tctataccga tgtggattgc gcatactttg tgaacagaaa 4141 gtgatagcgt tgatgattct tcattggtca gaaaattatg aacggtttct tctattttgt 4201 ctctatatac tacgtatagg aaatgtttac attttcgtat tgttttcgat tcactctatg 4261 aatagttctt actacaattt ttttgtctaa agagtaatac tagagataaa cataaaaaat 4321 gtagaggtcg agtttagatg caagttcaag gagcgaaagg tggatgggta ggttatatag 4381 ggatatagca cagagatata tagcaaagag atacttttga gcaatgtttg tggaagcggt 4441 attcgcaata ttttagtagc tcgttacagt ccggtgcgtt tttggttttt tgaaagtgcg 4501 tcatcagagc gcttttggtt ttcaaaagcg ctctgaagtt cctatacttt ctagctagag 4561 aataggaact tcccgagcgg ccgcgtgtta caagcttgac tccagcgtaa ctggactgca 4621 atcaactcac tggctcacct tcacgggtgg gcctttcttc ggtagaaaat caaaggatct 4681 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta 4741 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgag gtaactggct 4801 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact 4861 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg 4921 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata 4981 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga 5041 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag 5101 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg 5161 agcttccagg ggtaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac 5221 ttgagcatcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca 5281 acgcagaaag gcccacccga aggtgagcca ggtgattaca tttgggccct catcagaggt 5341 tttcaccgtc atcaccgaaa cgcgcgaggc agctgcggta aagctcatca gcgtggtcgt 5401 gaagcgattc acagatgtct gcctgttcat ccgcgtccag ctcgttgagt ttctccagaa 5461 gcgttaatgt ctggcttctg ataaagcggg ccatgttaag ggcggttttt tcctgtttgg 5521 tcacttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat 5581 ccatagttgc ctgactcccc gtcgtgtaga taactacgat gcgggagggc ttaccatctg 5641 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa 5701 taaaccagcc agccgggagg gccgagcgca gaagtgatcc tgcaacttta tccgcctcca 5761 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc 5821 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt 5881 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa 5941 aagcggttag ctccttcggt cctccgatcg ttgccagaag taagttggcc gcagtgttat 6001 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtgagatgct 6061 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga 6121 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag 6181 tgctcatcat tggaaaacgt tcttcggggc gtaaactctc aaggatctta ccgctgttga 6241 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca 6301 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg 6361 cgacacggaa atgttgaata ctcattttag cttccttagc tcctgaaaat ctcgataact 6421 caaaaaatac gcccggtagt gatcttattt cattatggtg aaagttggaa cctcttacgt 6481 gccgatcaag tcaaaagcct ccggtcggag gcttttgact ttctgctatg gaggtcaggt 6541 atgatttaaa tggtcagtta ga //