LOCUS Exported 6604 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Yeast expression plasmid for SARS-CoV-2 helicase. ACCESSION . VERSION . KEYWORDS pGBW-m4046248 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6604) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6604) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6604 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 55..462 /gene="S. cerevisiae TEF1" /label=TEF1 promoter /note="promoter for EF-1-alpha" CDS 2276..2293 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 2294..2359 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" primer_bind complement(2404..2422) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 2773..3576 /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 3659..4641 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 4719..4748 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 4770..5357 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 5369..5396 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(5420..5611) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(5612..6472) /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 6235..6254 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6473..6575) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 6576..3 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" ORIGIN 1 actttctgct atggaggtca ggtatgattt aaatggtcag ttagagcggc cgccccacac 61 accatagctt caaaatgttt ctactccttt tttactcttc cagattttct cggactccgc 121 gcatcgccgt accacttcaa aacacccaag cacagcatac taaatttccc ctctttcttc 181 ctctagggtg tcgttaatta cccgtactaa aggtttggaa aagaaaaaag agaccgcctc 241 gtttcttttt cttcgtcgaa aaaggcaata aaaattttta tcacgtttct ttttcttgaa 301 aatttttttt tttgattttt ttctctttcg atgacctccc attgatattt aagttaataa 361 acggtcttca atttctcaag tttcagtttc atttttcttg ttctattaca acttttttta 421 cttcttgctc attagaaaga aagcatagca atctaatcta agttttaatt acaaaatggt 481 tggggcttgt gttctttgca attcacagac ttcattaaga tgtggtgctt gcatacgtag 541 accattctta tgttgtaaat gctgttacga ccatgtcata tcaacatcac ataaattagt 601 cttgtctgtt aatccgtatg tttgcaatgc tccaggttgt gatgtcacag atgtgactca 661 actttactta ggaggtatga gctattattg taaatcacat aaaccaccca ttagttttcc 721 attgtgtgct aatggacaag tttttggttt atataaaaat acatgtgttg gtagcgataa 781 tgttactgac tttaatgcaa ttgcaacatg tgactggaca aatgctggtg attacatttt 841 agctaacacc tgtactgaaa gactcaagct ttttgcagca gaaacgctca aagctactga 901 ggagacattt aaactgtctt atggtattgc tactgtacgt gaagtgctgt ctgacagaga 961 attacatctt tcatgggaag ttggtaaacc tagaccacca cttaaccgaa attatgtctt 1021 tactggttat cgtgtaacta aaaacagtaa agtacaaata ggagagtaca cctttgaaaa 1081 aggtgactat ggtgatgctg ttgtttaccg aggtacaaca acttacaaat taaatgttgg 1141 tgattatttt gtgctgacat cacatacagt aatgccatta agtgcaccta cactagtgcc 1201 acaagagcac tatgttagaa ttactggctt atacccaaca ctcaatatct cagatgagtt 1261 ttctagcaat gttgcaaatt atcaaaaggt tggtatgcaa aagtattcta cactccaggg 1321 accacctggt actggtaaga gtcattttgc tattggccta gctctctact acccttctgc 1381 tcgcatagtg tatacagctt gctctcatgc cgctgttgat gcactatgtg agaaggcatt 1441 aaaatatttg cctatagata aatgtagtag aattatacct gcacgtgctc gtgtagagtg 1501 ttttgataaa ttcaaagtga attcaacatt agaacagtat gtcttttgta ctgtaaatgc 1561 attgcctgag acgacagcag atatagttgt ctttgatgaa atttcaatgg ccacaaatta 1621 tgatttgagt gttgtcaatg ccagattacg tgctaagcac tatgtgtaca ttggcgaccc 1681 tgctcaatta cctgcaccac gcacattgct aactaagggc acactagaac cagaatattt 1741 caattcagtg tgtagactta tgaaaactat aggtccagac atgttcctcg gaacttgtcg 1801 gcgttgtcct gctgaaattg ttgacactgt gagtgctttg gtttatgata ataagcttaa 1861 agcacataaa gacaaatcag ctcaatgctt taaaatgttt tataagggtg ttatcacgca 1921 tgatgtttca tctgcaatta acaggccaca aataggcgtg gtaagagaat tccttacacg 1981 taaccctgct tggagaaaag ctgtctttat ttcaccttat aattcacaga atgctgtagc 2041 ctcaaagatt ttgggactac caactcaaac tgttgattca tcacagggct cagaatatga 2101 ctatgtcata ttcactcaaa ccactgaaac agctcactct tgtaatgtaa acagatttaa 2161 tgttgctatt accagagcaa aagtaggcat actttgcata atgtctgata gagaccttta 2221 tgacaagttg caatttacaa gtcttgaaat tccacgtagg aatgtggcaa ctttactcgt 2281 gccgagaggt tctgattata aagatcatga tggtgattat aaagatcatg atattgatta 2341 taaagatgat gatgataaat aaacaggccc cttttccttt gtcgatatca tgtaattagt 2401 tatgtcacgc ttacattcac gccctcctcc cacatccgct ctaaccgaaa aggaaggagt 2461 tagacaacct gaagtctagg tccctattta ttttttttaa tagttatgtt agtattaaga 2521 acgttattta tatttcaaat ttttcttttt tttctgtaca aagcggccgc cgaaccacta 2581 gtgcactgca gtacacggtt tccttgaaat ttttttgatt cggtaatctc cgaacagaag 2641 gaagaacgaa ggaaggagca cagacttaga ttggtatata tacgcatatg tagtgttgaa 2701 gaaacatgaa attgcccagt attcttaacc caactgcaca gaacaaaaac gtgcaggaaa 2761 cgaagataaa tcatgtcgaa agctacatat aaggaacgtg ctgctactca tcctagtcct 2821 gttgctgcca agctatttaa tatcatgcac gaaaagcaaa caaacttgtg tgcttcattg 2881 gatgttcgta ccaccaagga attactggag ttagttgaag cattaggtcc caaaatttgt 2941 ttactaaaaa cacatgtgga tatcttgact gatttttcca tggagggcac agttaagccg 3001 ctaaaggcat tatccgccaa gtacaatttt ttactcttcg aggacagaaa atttgctgac 3061 attggtaata cagtcaaatt gcagtactct gcgggtgtat acagaatagc agaatgggca 3121 gacattacga atgcacacgg tgtggtgggc ccaggtattg ttagcggttt gaagcaggcg 3181 gcagaagaag taacaaagga acctagaggc cttttgatgt tagcagaatt gtcatgcaag 3241 ggctccctat ctactggaga atatactaag ggtactgttg acattgcgaa gagcgacaaa 3301 gattttgtta tcggctttat tgctcaaaga gacatgggtg gaagagatga aggttacgat 3361 tggttgatta tgacacccgg tgtgggttta gatgacaagg gagatgcatt gggtcaacag 3421 tatagaaccg tggatgatgt ggtttctaca ggatctgaca ttattattgt tggaagagga 3481 ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa agcaggctgg 3541 gaagcatatt tgagaagatg cggccagcaa aactaaaaaa ctgtattata agtaaatgca 3601 tgtatactaa actcacaaat tagagcttca atttaattat atcagttatt acccgagtaa 3661 cgaagcatct gtgcttcatt ttgtagaaca aaaatgcaac gcgagagcgc taatttttca 3721 aacaaagaat ctgagctgca tttttacaga acagaaatgc aacgcgaaag cgctatttta 3781 ccaacgaaga atctgtgctt catttttgta aaacaaaaat gcaacgcgag agcgctaatt 3841 tttcaaacaa agaatctgag ctgcattttt acagaacaga aatgcaacgc gagagcgcta 3901 ttttaccaac aaagaatcta tacttctttt ttgttctaca aaaatgcatc ccgagagcgc 3961 tatttttcta acaaagcatc ttagattact ttttttctcc tttgtgcgct ctataatgca 4021 gtctcttgat aactttttgc actgtaggtc cgttaaggtt agaagaaggc tactttggtg 4081 tctattttct cttccataaa aaaagcctga ctccacttcc cgcgtttact gattactagc 4141 gaagctgcgg gtgcattttt tcaagataaa ggcatccccg attatattct ataccgatgt 4201 ggattgcgca tactttgtga acagaaagtg atagcgttga tgattcttca ttggtcagaa 4261 aattatgaac ggtttcttct attttgtctc tatatactac gtataggaaa tgtttacatt 4321 ttcgtattgt tttcgattca ctctatgaat agttcttact acaatttttt tgtctaaaga 4381 gtaatactag agataaacat aaaaaatgta gaggtcgagt ttagatgcaa gttcaaggag 4441 cgaaaggtgg atgggtaggt tatataggga tatagcacag agatatatag caaagagata 4501 cttttgagca atgtttgtgg aagcggtatt cgcaatattt tagtagctcg ttacagtccg 4561 gtgcgttttt ggttttttga aagtgcgtca tcagagcgct tttggttttc aaaagcgctc 4621 tgaagttcct atactttcta gctagagaat aggaacttcc cgagcggccg cgtgttacaa 4681 gcttgactcc agcgtaactg gactgcaatc aactcactgg ctcaccttca cgggtgggcc 4741 tttcttcggt agaaaatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc 4801 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta 4861 ccaactcttt ttccgaggta actggcttca gcagagcgca gataccaaat actgttcttc 4921 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg 4981 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt 5041 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt 5101 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc 5161 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca 5221 gggtcggaac aggagagcgc acgagggagc ttccaggggt aaacgcctgg tatctttata 5281 gtcctgtcgg gtttcgccac ctctgacttg agcatcgatt tttgtgatgc tcgtcagggg 5341 ggcggagcct atggaaaaac gccagcaacg cagaaaggcc cacccgaagg tgagccaggt 5401 gattacattt gggccctcat cagaggtttt caccgtcatc accgaaacgc gcgaggcagc 5461 tgcggtaaag ctcatcagcg tggtcgtgaa gcgattcaca gatgtctgcc tgttcatccg 5521 cgtccagctc gttgagtttc tccagaagcg ttaatgtctg gcttctgata aagcgggcca 5581 tgttaagggc ggttttttcc tgtttggtca cttaccaatg cttaatcagt gaggcaccta 5641 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa 5701 ctacgatgcg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac 5761 gctcaccggc tccagattta tcagcaataa accagccagc cgggagggcc gagcgcagaa 5821 gtgatcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag 5881 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg 5941 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag 6001 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg 6061 ccagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc 6121 ttactgtcat gccatccgtg agatgctttt ctgtgactgg tgagtactca accaagtcat 6181 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata 6241 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgta 6301 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca 6361 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc 6421 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc attttagctt 6481 ccttagctcc tgaaaatctc gataactcaa aaaatacgcc cggtagtgat cttatttcat 6541 tatggtgaaa gttggaacct cttacgtgcc gatcaagtca aaagcctccg gtcggaggct 6601 tttg //