LOCUS Exported 5677 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Yeast Expression plasmid for SARS-CoV-2 3C-like proteinase. ACCESSION . VERSION . KEYWORDS pGBW-m4046345 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5677) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5677) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5677 /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 1352..1369 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 1370..1387 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(1432..1450) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 1801..2604 /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 2687..3669 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 3747..3776 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 3798..4385 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 4397..4424 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(4448..4639) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4640..5500) /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 5263..5282 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5501..5603) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 5604..5635 /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 atgtcgggtt tcagaaaaat ggcctttcct agtggaaagg tcgagggctg 481 tatggtgcaa gtaacatgcg gtacgaccac tttgaacggt ttatggcttg acgatgttgt 541 ttactgtcca cgtcatgtca tctgtacttc agaagatatg ttgaatccaa actatgaaga 601 cctactgatt cgcaagtcta accacaattt cttagttcag gctggtaacg ttcaattgag 661 agttataggt cacagcatgc aaaattgtgt cttgaagtta aaagttgata ccgcaaaccc 721 gaagactcca aagtacaaat ttgtcagaat ccaacccggt caaaccttct ccgtcttggc 781 ttgttataac gggtccccat ctggtgtata ccagtgcgct atgaggccaa acttcactat 841 taagggttct tttttgaatg gttcttgtgg ctccgttggt tttaatatcg actacgattg 901 cgtcagtttc tgttacatgc atcacatgga attgcctaca ggtgttcacg ccggaaccga 961 cctagaaggg aacttctatg gtccattcgt tgatcggcaa actgctcaag ctgcgggtac 1021 ggataccaca attactgtaa atgtgcttgc ctggttgtac gctgctgtta taaacggtga 1081 ccgatggttt ttaaacagat tcactaccac attgaatgac tttaatctgg tcgctatgaa 1141 atataactac gaaccactca ctcaagatca tgtcgatatt ttaggtcctt tgtctgcaca 1201 aactggcatc gctgttttgg acatgtgtgc gtcccttaag gaactactcc aaaacggtat 1261 gaacgggaga acgattttgg gttcagccct attggaggac gaatttaccc cattcgatgt 1321 ggttagacag tgctctggag taaccttcca actcgtgccg agaggttctc accatcatca 1381 ccatcattaa acaggcccct tttcctttgt cgatatcatg taattagtta tgtcacgctt 1441 acattcacgc cctcctccca catccgctct aaccgaaaag gaaggagtta gacaacctga 1501 agtctaggtc cctatttatt ttttttaata gttatgttag tattaagaac gttatttata 1561 tttcaaattt ttcttttttt tctgtacaaa gcggccgccg aaccactagt gcactgcagt 1621 acacggtttc cttgaaattt ttttgattcg gtaatctccg aacagaagga agaacgaagg 1681 aaggagcaca gacttagatt ggtatatata cgcatatgta gtgttgaaga aacatgaaat 1741 tgcccagtat tcttaaccca actgcacaga acaaaaacgt gcaggaaacg aagataaatc 1801 atgtcgaaag ctacatataa ggaacgtgct gctactcatc ctagtcctgt tgctgccaag 1861 ctatttaata tcatgcacga aaagcaaaca aacttgtgtg cttcattgga tgttcgtacc 1921 accaaggaat tactggagtt agttgaagca ttaggtccca aaatttgttt actaaaaaca 1981 catgtggata tcttgactga tttttccatg gagggcacag ttaagccgct aaaggcatta 2041 tccgccaagt acaatttttt actcttcgag gacagaaaat ttgctgacat tggtaataca 2101 gtcaaattgc agtactctgc gggtgtatac agaatagcag aatgggcaga cattacgaat 2161 gcacacggtg tggtgggccc aggtattgtt agcggtttga agcaggcggc agaagaagta 2221 acaaaggaac ctagaggcct tttgatgtta gcagaattgt catgcaaggg ctccctatct 2281 actggagaat atactaaggg tactgttgac attgcgaaga gcgacaaaga ttttgttatc 2341 ggctttattg ctcaaagaga catgggtgga agagatgaag gttacgattg gttgattatg 2401 acacccggtg tgggtttaga tgacaaggga gatgcattgg gtcaacagta tagaaccgtg 2461 gatgatgtgg tttctacagg atctgacatt attattgttg gaagaggact atttgcaaag 2521 ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag caggctggga agcatatttg 2581 agaagatgcg gccagcaaaa ctaaaaaact gtattataag taaatgcatg tatactaaac 2641 tcacaaatta gagcttcaat ttaattatat cagttattac ccgagtaacg aagcatctgt 2701 gcttcatttt gtagaacaaa aatgcaacgc gagagcgcta atttttcaaa caaagaatct 2761 gagctgcatt tttacagaac agaaatgcaa cgcgaaagcg ctattttacc aacgaagaat 2821 ctgtgcttca tttttgtaaa acaaaaatgc aacgcgagag cgctaatttt tcaaacaaag 2881 aatctgagct gcatttttac agaacagaaa tgcaacgcga gagcgctatt ttaccaacaa 2941 agaatctata cttctttttt gttctacaaa aatgcatccc gagagcgcta tttttctaac 3001 aaagcatctt agattacttt ttttctcctt tgtgcgctct ataatgcagt ctcttgataa 3061 ctttttgcac tgtaggtccg ttaaggttag aagaaggcta ctttggtgtc tattttctct 3121 tccataaaaa aagcctgact ccacttcccg cgtttactga ttactagcga agctgcgggt 3181 gcattttttc aagataaagg catccccgat tatattctat accgatgtgg attgcgcata 3241 ctttgtgaac agaaagtgat agcgttgatg attcttcatt ggtcagaaaa ttatgaacgg 3301 tttcttctat tttgtctcta tatactacgt ataggaaatg tttacatttt cgtattgttt 3361 tcgattcact ctatgaatag ttcttactac aatttttttg tctaaagagt aatactagag 3421 ataaacataa aaaatgtaga ggtcgagttt agatgcaagt tcaaggagcg aaaggtggat 3481 gggtaggtta tatagggata tagcacagag atatatagca aagagatact tttgagcaat 3541 gtttgtggaa gcggtattcg caatatttta gtagctcgtt acagtccggt gcgtttttgg 3601 ttttttgaaa gtgcgtcatc agagcgcttt tggttttcaa aagcgctctg aagttcctat 3661 actttctagc tagagaatag gaacttcccg agcggccgcg tgttacaagc ttgactccag 3721 cgtaactgga ctgcaatcaa ctcactggct caccttcacg ggtgggcctt tcttcggtag 3781 aaaatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca 3841 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt 3901 ccgaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt 3961 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc 4021 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac 4081 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca 4141 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg 4201 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag 4261 gagagcgcac gagggagctt ccaggggtaa acgcctggta tctttatagt cctgtcgggt 4321 ttcgccacct ctgacttgag catcgatttt tgtgatgctc gtcagggggg cggagcctat 4381 ggaaaaacgc cagcaacgca gaaaggccca cccgaaggtg agccaggtga ttacatttgg 4441 gccctcatca gaggttttca ccgtcatcac cgaaacgcgc gaggcagctg cggtaaagct 4501 catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg ttcatccgcg tccagctcgt 4561 tgagtttctc cagaagcgtt aatgtctggc ttctgataaa gcgggccatg ttaagggcgg 4621 ttttttcctg tttggtcact taccaatgct taatcagtga ggcacctatc tcagcgatct 4681 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatgcggg 4741 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc 4801 cagatttatc agcaataaac cagccagccg ggagggccga gcgcagaagt gatcctgcaa 4861 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc 4921 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt 4981 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc 5041 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgcc agaagtaagt 5101 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc 5161 catccgtgag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt 5221 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata 5281 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgtaaa ctctcaagga 5341 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag 5401 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa 5461 aaaagggaat aagggcgaca cggaaatgtt gaatactcat tttagcttcc ttagctcctg 5521 aaaatctcga taactcaaaa aatacgcccg gtagtgatct tatttcatta tggtgaaagt 5581 tggaacctct tacgtgccga tcaagtcaaa agcctccggt cggaggcttt tgactttctg 5641 ctatggaggt caggtatgat ttaaatggtc agttaga //