LOCUS Exported 5706 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Yeast Expression plasmid for SARS-CoV-2 2'-O-ribose methyltransferase. ACCESSION . VERSION . KEYWORDS pGBW-m4046556 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5706) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5706) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5706 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 25..466 /gene="S. cerevisiae GAL1" /label=GAL1 promoter /note="inducible promoter, regulated by Gal4" protein_bind 25..142 /label=UAS /bound_moiety="Gal4" /note="upstream activating sequence mediating Gal4-dependent induction" primer_bind 437..460 /label=GAL1 /note="S. cerevisiae GAL1 promoter, forward primer" CDS 1375..1392 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 1393..1416 /codon_start=1 /product="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" /label=Strep-Tag II /translation="WSHPQFEK" primer_bind complement(1461..1479) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 1830..2633 /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 2716..3698 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 3776..3805 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 3827..4414 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 4426..4453 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(4477..4668) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4669..5529) /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 5292..5311 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5530..5632) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 5633..5664 /label=tonB terminator /note="bidirectional E. coli tonB-P14 transcription terminator" ORIGIN 1 gcggccgctt gctatattga agtacggatt agaagccgcc gagcgggcga cagccctccg 61 acggaagact ctcctccgtg cgtcctcgtc ttcaccggtc gcgttcctga aacgcagatg 121 tgcctcgcgc cgcactgctc cgaacaataa agattctaca atactagctt ttatggttat 181 gaagaggaaa aattggcagt aacctggccc cacaaacctt caaattaacg aatcaaatta 241 acaaccatag gatgataatg cgattagttt tttagcctta tttctggggt aattaatcag 301 cgaagcgatg atttttgatc tattaacaga tatataaatg gaaaagctgc ataaccactt 361 taactaatac tttcaacatt ttcagtttgt attacttctt attcaaatgt cataaaagta 421 tcaacaaaaa attgttaata tacctctata ctttaacgtc aaggagaaaa aactataatg 481 tcctctcaag cctggcagcc tggtgtggct atgcccaatt tgtataagat gcaaagaatg 541 cttctggaga aatgcgattt gcaaaactac ggagacagcg ctacattacc aaagggtatt 601 atgatgaacg tagcaaagta cacccaattg tgtcaatact taaatacgtt gactttggct 661 gttccgtata acatgcgtgt catacacttt ggggccggtt cagacaaagg cgtcgctcca 721 ggtaccgcag ttctaaggca atggctccca actggtactc tattggttga ttctgatctg 781 aacgacttcg tcagtgatgc tgactccact ttaatcggtg attgtgctac cgttcatacg 841 gcgaacaagt gggacttgat tatttcggat atgtacgacc caaagaccaa aaatgttaca 901 aaggaaaatg attctaaaga aggtttcttt acttatatct gtggtttcat ccagcaaaag 961 cttgccttgg gcggttctgt ggctattaag atcactgaac actcctggaa cgccgactta 1021 tacaaattga tgggtcattt cgcttggtgg accgcgtttg ttaccaacgt taacgctagt 1081 tcttcggaag cctttctcat cggttgtaat tatttaggaa agccacgcga acaaattgat 1141 ggttacgtca tgcacgctaa ctacattttc tggagaaata ctaaccctat acaattgtca 1201 agctatagtt tattcgacat gtcaaagttt ccattgaagt tgagaggtac agctgtcatg 1261 tccttgaagg aagggcagat caacgatatg atcttatctt tgctgtctaa aggtagactc 1321 attattagag aaaataatcg ggttgttatc tcttctgatg tgctagtcaa caacctcgtg 1381 ccgagaggtt cttggagcca tccgcagttt gaaaaataaa caggcccctt ttcctttgtc 1441 gatatcatgt aattagttat gtcacgctta cattcacgcc ctcctcccac atccgctcta 1501 accgaaaagg aaggagttag acaacctgaa gtctaggtcc ctatttattt tttttaatag 1561 ttatgttagt attaagaacg ttatttatat ttcaaatttt tctttttttt ctgtacaaag 1621 cggccgccga accactagtg cactgcagta cacggtttcc ttgaaatttt tttgattcgg 1681 taatctccga acagaaggaa gaacgaagga aggagcacag acttagattg gtatatatac 1741 gcatatgtag tgttgaagaa acatgaaatt gcccagtatt cttaacccaa ctgcacagaa 1801 caaaaacgtg caggaaacga agataaatca tgtcgaaagc tacatataag gaacgtgctg 1861 ctactcatcc tagtcctgtt gctgccaagc tatttaatat catgcacgaa aagcaaacaa 1921 acttgtgtgc ttcattggat gttcgtacca ccaaggaatt actggagtta gttgaagcat 1981 taggtcccaa aatttgttta ctaaaaacac atgtggatat cttgactgat ttttccatgg 2041 agggcacagt taagccgcta aaggcattat ccgccaagta caatttttta ctcttcgagg 2101 acagaaaatt tgctgacatt ggtaatacag tcaaattgca gtactctgcg ggtgtataca 2161 gaatagcaga atgggcagac attacgaatg cacacggtgt ggtgggccca ggtattgtta 2221 gcggtttgaa gcaggcggca gaagaagtaa caaaggaacc tagaggcctt ttgatgttag 2281 cagaattgtc atgcaagggc tccctatcta ctggagaata tactaagggt actgttgaca 2341 ttgcgaagag cgacaaagat tttgttatcg gctttattgc tcaaagagac atgggtggaa 2401 gagatgaagg ttacgattgg ttgattatga cacccggtgt gggtttagat gacaagggag 2461 atgcattggg tcaacagtat agaaccgtgg atgatgtggt ttctacagga tctgacatta 2521 ttattgttgg aagaggacta tttgcaaagg gaagggatgc taaggtagag ggtgaacgtt 2581 acagaaaagc aggctgggaa gcatatttga gaagatgcgg ccagcaaaac taaaaaactg 2641 tattataagt aaatgcatgt atactaaact cacaaattag agcttcaatt taattatatc 2701 agttattacc cgagtaacga agcatctgtg cttcattttg tagaacaaaa atgcaacgcg 2761 agagcgctaa tttttcaaac aaagaatctg agctgcattt ttacagaaca gaaatgcaac 2821 gcgaaagcgc tattttacca acgaagaatc tgtgcttcat ttttgtaaaa caaaaatgca 2881 acgcgagagc gctaattttt caaacaaaga atctgagctg catttttaca gaacagaaat 2941 gcaacgcgag agcgctattt taccaacaaa gaatctatac ttcttttttg ttctacaaaa 3001 atgcatcccg agagcgctat ttttctaaca aagcatctta gattactttt tttctccttt 3061 gtgcgctcta taatgcagtc tcttgataac tttttgcact gtaggtccgt taaggttaga 3121 agaaggctac tttggtgtct attttctctt ccataaaaaa agcctgactc cacttcccgc 3181 gtttactgat tactagcgaa gctgcgggtg cattttttca agataaaggc atccccgatt 3241 atattctata ccgatgtgga ttgcgcatac tttgtgaaca gaaagtgata gcgttgatga 3301 ttcttcattg gtcagaaaat tatgaacggt ttcttctatt ttgtctctat atactacgta 3361 taggaaatgt ttacattttc gtattgtttt cgattcactc tatgaatagt tcttactaca 3421 atttttttgt ctaaagagta atactagaga taaacataaa aaatgtagag gtcgagttta 3481 gatgcaagtt caaggagcga aaggtggatg ggtaggttat atagggatat agcacagaga 3541 tatatagcaa agagatactt ttgagcaatg tttgtggaag cggtattcgc aatattttag 3601 tagctcgtta cagtccggtg cgtttttggt tttttgaaag tgcgtcatca gagcgctttt 3661 ggttttcaaa agcgctctga agttcctata ctttctagct agagaatagg aacttcccga 3721 gcggccgcgt gttacaagct tgactccagc gtaactggac tgcaatcaac tcactggctc 3781 accttcacgg gtgggccttt cttcggtaga aaatcaaagg atcttcttga gatccttttt 3841 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt 3901 tgccggatca agagctacca actctttttc cgaggtaact ggcttcagca gagcgcagat 3961 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc 4021 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa 4081 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg 4141 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag 4201 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag 4261 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc caggggtaaa 4321 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc atcgattttt 4381 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcag aaaggcccac 4441 ccgaaggtga gccaggtgat tacatttggg ccctcatcag aggttttcac cgtcatcacc 4501 gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg attcacagat 4561 gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta atgtctggct 4621 tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactt accaatgctt 4681 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact 4741 ccccgtcgtg tagataacta cgatgcggga gggcttacca tctggcccca gtgctgcaat 4801 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg 4861 gagggccgag cgcagaagtg atcctgcaac tttatccgcc tccatccagt ctattaattg 4921 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat 4981 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc 5041 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt 5101 cggtcctccg atcgttgcca gaagtaagtt ggccgcagtg ttatcactca tggttatggc 5161 agcactgcat aattctctta ctgtcatgcc atccgtgaga tgcttttctg tgactggtga 5221 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc 5281 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa 5341 acgttcttcg gggcgtaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta 5401 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg 5461 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg 5521 aatactcatt ttagcttcct tagctcctga aaatctcgat aactcaaaaa atacgcccgg 5581 tagtgatctt atttcattat ggtgaaagtt ggaacctctt acgtgccgat caagtcaaaa 5641 gcctccggtc ggaggctttt gactttctgc tatggaggtc aggtatgatt taaatggtca 5701 gttaga //