LOCUS Exported 5671 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Yeast Expression plasmid for SARS-CoV-2 3C-like proteinase. ACCESSION . VERSION . KEYWORDS pGBW-m4046440 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5671) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5671) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5671 /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 1346..1363 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 1364..1381 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(1426..1444) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 1795..2598 /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 2681..3663 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 3741..3770 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 3792..4379 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 4391..4418 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(4442..4633) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4634..5494) /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 5257..5276 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5495..5597) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 5598..5629 /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 atgggtttta gaaaaatggc attcccatct ggtaaagttg agggttgtat 481 ggtacaagta acttgtggta caactacact taacggtctt tggcttgatg acgtagttta 541 ctgtccaaga catgtgatct gcacctctga agacatgctt aaccctaatt atgaagattt 601 actcattcgt aagtctaatc ataatttctt ggtacaggct ggtaatgttc aactcagggt 661 tattggacat tctatgcaaa attgtgtact taagcttaag gttgatacag ccaatcctaa 721 gacacctaag tataagtttg ttcgcattca accaggacag actttttcag tgttagcttg 781 ttacaatggt tcaccatctg gtgtttacca atgtgctatg aggcccaatt tcactattaa 841 gggttcattc cttaatggtt catgtggtag tgttggtttt aacatagatt atgactgtgt 901 ctctttttgt tacatgcacc atatggaatt accaactgga gttcatgctg gcacagactt 961 agaaggtaac ttttatggac cttttgttga caggcaaaca gcacaagcag ctggtacgga 1021 cacaactatt acagttaatg ttttagcttg gttgtacgct gctgttataa atggagacag 1081 gtggtttctc aatcgattta ccacaactct taatgacttt aaccttgtgg ctatgaagta 1141 caattatgaa cctctaacac aagaccatgt tgacatacta ggacctcttt ctgctcaaac 1201 tggaattgcc gttttagata tgtgtgcttc attaaaagaa ttactgcaaa atggtatgaa 1261 tggacgtacc atattgggta gtgctttatt agaagatgaa tttacacctt ttgatgttgt 1321 tagacaatgc tcaggtgtta ctttcctcgt gccgagaggt tctcaccatc atcaccatca 1381 ttaaacaggc cccttttcct ttgtcgatat catgtaatta gttatgtcac gcttacattc 1441 acgccctcct cccacatccg ctctaaccga aaaggaagga gttagacaac ctgaagtcta 1501 ggtccctatt tatttttttt aatagttatg ttagtattaa gaacgttatt tatatttcaa 1561 atttttcttt tttttctgta caaagcggcc gccgaaccac tagtgcactg cagtacacgg 1621 tttccttgaa atttttttga ttcggtaatc tccgaacaga aggaagaacg aaggaaggag 1681 cacagactta gattggtata tatacgcata tgtagtgttg aagaaacatg aaattgccca 1741 gtattcttaa cccaactgca cagaacaaaa acgtgcagga aacgaagata aatcatgtcg 1801 aaagctacat ataaggaacg tgctgctact catcctagtc ctgttgctgc caagctattt 1861 aatatcatgc acgaaaagca aacaaacttg tgtgcttcat tggatgttcg taccaccaag 1921 gaattactgg agttagttga agcattaggt cccaaaattt gtttactaaa aacacatgtg 1981 gatatcttga ctgatttttc catggagggc acagttaagc cgctaaaggc attatccgcc 2041 aagtacaatt ttttactctt cgaggacaga aaatttgctg acattggtaa tacagtcaaa 2101 ttgcagtact ctgcgggtgt atacagaata gcagaatggg cagacattac gaatgcacac 2161 ggtgtggtgg gcccaggtat tgttagcggt ttgaagcagg cggcagaaga agtaacaaag 2221 gaacctagag gccttttgat gttagcagaa ttgtcatgca agggctccct atctactgga 2281 gaatatacta agggtactgt tgacattgcg aagagcgaca aagattttgt tatcggcttt 2341 attgctcaaa gagacatggg tggaagagat gaaggttacg attggttgat tatgacaccc 2401 ggtgtgggtt tagatgacaa gggagatgca ttgggtcaac agtatagaac cgtggatgat 2461 gtggtttcta caggatctga cattattatt gttggaagag gactatttgc aaagggaagg 2521 gatgctaagg tagagggtga acgttacaga aaagcaggct gggaagcata tttgagaaga 2581 tgcggccagc aaaactaaaa aactgtatta taagtaaatg catgtatact aaactcacaa 2641 attagagctt caatttaatt atatcagtta ttacccgagt aacgaagcat ctgtgcttca 2701 ttttgtagaa caaaaatgca acgcgagagc gctaattttt caaacaaaga atctgagctg 2761 catttttaca gaacagaaat gcaacgcgaa agcgctattt taccaacgaa gaatctgtgc 2821 ttcatttttg taaaacaaaa atgcaacgcg agagcgctaa tttttcaaac aaagaatctg 2881 agctgcattt ttacagaaca gaaatgcaac gcgagagcgc tattttacca acaaagaatc 2941 tatacttctt ttttgttcta caaaaatgca tcccgagagc gctatttttc taacaaagca 3001 tcttagatta ctttttttct cctttgtgcg ctctataatg cagtctcttg ataacttttt 3061 gcactgtagg tccgttaagg ttagaagaag gctactttgg tgtctatttt ctcttccata 3121 aaaaaagcct gactccactt cccgcgttta ctgattacta gcgaagctgc gggtgcattt 3181 tttcaagata aaggcatccc cgattatatt ctataccgat gtggattgcg catactttgt 3241 gaacagaaag tgatagcgtt gatgattctt cattggtcag aaaattatga acggtttctt 3301 ctattttgtc tctatatact acgtatagga aatgtttaca ttttcgtatt gttttcgatt 3361 cactctatga atagttctta ctacaatttt tttgtctaaa gagtaatact agagataaac 3421 ataaaaaatg tagaggtcga gtttagatgc aagttcaagg agcgaaaggt ggatgggtag 3481 gttatatagg gatatagcac agagatatat agcaaagaga tacttttgag caatgtttgt 3541 ggaagcggta ttcgcaatat tttagtagct cgttacagtc cggtgcgttt ttggtttttt 3601 gaaagtgcgt catcagagcg cttttggttt tcaaaagcgc tctgaagttc ctatactttc 3661 tagctagaga ataggaactt cccgagcggc cgcgtgttac aagcttgact ccagcgtaac 3721 tggactgcaa tcaactcact ggctcacctt cacgggtggg cctttcttcg gtagaaaatc 3781 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa 3841 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgagg 3901 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag 3961 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac 4021 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt 4081 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg 4141 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc 4201 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc 4261 gcacgaggga gcttccaggg gtaaacgcct ggtatcttta tagtcctgtc gggtttcgcc 4321 acctctgact tgagcatcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa 4381 acgccagcaa cgcagaaagg cccacccgaa ggtgagccag gtgattacat ttgggccctc 4441 atcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa agctcatcag 4501 cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc tcgttgagtt 4561 tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg gcggtttttt 4621 cctgtttggt cacttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat 4681 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgatg cgggagggct 4741 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt 4801 tatcagcaat aaaccagcca gccgggaggg ccgagcgcag aagtgatcct gcaactttat 4861 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta 4921 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg 4981 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt 5041 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgccagaagt aagttggccg 5101 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg 5161 tgagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc 5221 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa 5281 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg taaactctca aggatcttac 5341 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt 5401 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg 5461 gaataagggc gacacggaaa tgttgaatac tcattttagc ttccttagct cctgaaaatc 5521 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac 5581 ctcttacgtg ccgatcaagt caaaagcctc cggtcggagg cttttgactt tctgctatgg 5641 aggtcaggta tgatttaaat ggtcagttag a //