LOCUS Exported 5695 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Yeast Expression plasmid for SARS-CoV-2 2'-O-ribose methyltransferase. ACCESSION . VERSION . KEYWORDS pGBW-m4046481 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5695) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5695) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5695 /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 1322..1339 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 1340..1405 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" primer_bind complement(1450..1468) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 1819..2622 /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 2705..3687 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 3765..3794 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 3816..4403 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 4415..4442 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(4466..4657) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4658..5518) /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 5281..5300 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(5519..5621) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 5622..5653 /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 atgagtcaag cgtggcaacc gggtgttgct atgcctaatc tttacaaaat 481 gcaaagaatg ctattagaaa agtgtgacct tcaaaattat ggtgatagtg caacattacc 541 taaaggcata atgatgaatg tcgcaaaata tactcaactg tgtcaatatt taaacacatt 601 aacattagct gtaccctata atatgagagt tatacatttt ggtgctggtt ctgataaagg 661 agttgcacca ggtacagctg ttttaagaca gtggttgcct acgggtacgc tgcttgtcga 721 ttcagatctt aatgactttg tctctgatgc agattcaact ttgattggtg attgtgcaac 781 tgtacataca gctaataaat gggatctcat tattagtgat atgtacgacc ctaagactaa 841 aaatgttaca aaagaaaatg actctaaaga gggttttttc acttacattt gtgggtttat 901 acaacaaaag ctagctcttg gaggttccgt ggctataaag ataacagaac attcttggaa 961 tgctgatctt tataagctca tgggacactt cgcatggtgg acagcctttg ttactaatgt 1021 gaatgcgtca tcatctgaag catttttaat tggatgtaat tatcttggca aaccacgcga 1081 acaaatagat ggttatgtca tgcatgcaaa ttacatattt tggaggaata caaatccaat 1141 tcagttgtct tcctattctt tatttgacat gagtaaattt ccccttaaat taaggggtac 1201 tgctgttatg tctttaaaag aaggtcaaat caatgatatg attttatctc ttcttagtaa 1261 aggtagactt ataattagag aaaacaacag agttgttatt tctagtgatg ttcttgttaa 1321 cctcgtgccg agaggttctg attataaaga tcatgatggt gattataaag atcatgatat 1381 tgattataaa gatgatgatg ataaataaac aggccccttt tcctttgtcg atatcatgta 1441 attagttatg tcacgcttac attcacgccc tcctcccaca tccgctctaa ccgaaaagga 1501 aggagttaga caacctgaag tctaggtccc tatttatttt ttttaatagt tatgttagta 1561 ttaagaacgt tatttatatt tcaaattttt cttttttttc tgtacaaagc ggccgccgaa 1621 ccactagtgc actgcagtac acggtttcct tgaaattttt ttgattcggt aatctccgaa 1681 cagaaggaag aacgaaggaa ggagcacaga cttagattgg tatatatacg catatgtagt 1741 gttgaagaaa catgaaattg cccagtattc ttaacccaac tgcacagaac aaaaacgtgc 1801 aggaaacgaa gataaatcat gtcgaaagct acatataagg aacgtgctgc tactcatcct 1861 agtcctgttg ctgccaagct atttaatatc atgcacgaaa agcaaacaaa cttgtgtgct 1921 tcattggatg ttcgtaccac caaggaatta ctggagttag ttgaagcatt aggtcccaaa 1981 atttgtttac taaaaacaca tgtggatatc ttgactgatt tttccatgga gggcacagtt 2041 aagccgctaa aggcattatc cgccaagtac aattttttac tcttcgagga cagaaaattt 2101 gctgacattg gtaatacagt caaattgcag tactctgcgg gtgtatacag aatagcagaa 2161 tgggcagaca ttacgaatgc acacggtgtg gtgggcccag gtattgttag cggtttgaag 2221 caggcggcag aagaagtaac aaaggaacct agaggccttt tgatgttagc agaattgtca 2281 tgcaagggct ccctatctac tggagaatat actaagggta ctgttgacat tgcgaagagc 2341 gacaaagatt ttgttatcgg ctttattgct caaagagaca tgggtggaag agatgaaggt 2401 tacgattggt tgattatgac acccggtgtg ggtttagatg acaagggaga tgcattgggt 2461 caacagtata gaaccgtgga tgatgtggtt tctacaggat ctgacattat tattgttgga 2521 agaggactat ttgcaaaggg aagggatgct aaggtagagg gtgaacgtta cagaaaagca 2581 ggctgggaag catatttgag aagatgcggc cagcaaaact aaaaaactgt attataagta 2641 aatgcatgta tactaaactc acaaattaga gcttcaattt aattatatca gttattaccc 2701 gagtaacgaa gcatctgtgc ttcattttgt agaacaaaaa tgcaacgcga gagcgctaat 2761 ttttcaaaca aagaatctga gctgcatttt tacagaacag aaatgcaacg cgaaagcgct 2821 attttaccaa cgaagaatct gtgcttcatt tttgtaaaac aaaaatgcaa cgcgagagcg 2881 ctaatttttc aaacaaagaa tctgagctgc atttttacag aacagaaatg caacgcgaga 2941 gcgctatttt accaacaaag aatctatact tcttttttgt tctacaaaaa tgcatcccga 3001 gagcgctatt tttctaacaa agcatcttag attacttttt ttctcctttg tgcgctctat 3061 aatgcagtct cttgataact ttttgcactg taggtccgtt aaggttagaa gaaggctact 3121 ttggtgtcta ttttctcttc cataaaaaaa gcctgactcc acttcccgcg tttactgatt 3181 actagcgaag ctgcgggtgc attttttcaa gataaaggca tccccgatta tattctatac 3241 cgatgtggat tgcgcatact ttgtgaacag aaagtgatag cgttgatgat tcttcattgg 3301 tcagaaaatt atgaacggtt tcttctattt tgtctctata tactacgtat aggaaatgtt 3361 tacattttcg tattgttttc gattcactct atgaatagtt cttactacaa tttttttgtc 3421 taaagagtaa tactagagat aaacataaaa aatgtagagg tcgagtttag atgcaagttc 3481 aaggagcgaa aggtggatgg gtaggttata tagggatata gcacagagat atatagcaaa 3541 gagatacttt tgagcaatgt ttgtggaagc ggtattcgca atattttagt agctcgttac 3601 agtccggtgc gtttttggtt ttttgaaagt gcgtcatcag agcgcttttg gttttcaaaa 3661 gcgctctgaa gttcctatac tttctagcta gagaatagga acttcccgag cggccgcgtg 3721 ttacaagctt gactccagcg taactggact gcaatcaact cactggctca ccttcacggg 3781 tgggcctttc ttcggtagaa aatcaaagga tcttcttgag atcctttttt tctgcgcgta 3841 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa 3901 gagctaccaa ctctttttcc gaggtaactg gcttcagcag agcgcagata ccaaatactg 3961 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat 4021 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta 4081 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg 4141 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc 4201 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa 4261 gcggcagggt cggaacagga gagcgcacga gggagcttcc aggggtaaac gcctggtatc 4321 tttatagtcc tgtcgggttt cgccacctct gacttgagca tcgatttttg tgatgctcgt 4381 caggggggcg gagcctatgg aaaaacgcca gcaacgcaga aaggcccacc cgaaggtgag 4441 ccaggtgatt acatttgggc cctcatcaga ggttttcacc gtcatcaccg aaacgcgcga 4501 ggcagctgcg gtaaagctca tcagcgtggt cgtgaagcga ttcacagatg tctgcctgtt 4561 catccgcgtc cagctcgttg agtttctcca gaagcgttaa tgtctggctt ctgataaagc 4621 gggccatgtt aagggcggtt ttttcctgtt tggtcactta ccaatgctta atcagtgagg 4681 cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt 4741 agataactac gatgcgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag 4801 acccacgctc accggctcca gatttatcag caataaacca gccagccggg agggccgagc 4861 gcagaagtga tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag 4921 ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca 4981 tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa 5041 ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga 5101 tcgttgccag aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata 5161 attctcttac tgtcatgcca tccgtgagat gcttttctgt gactggtgag tactcaacca 5221 agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg 5281 ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg 5341 ggcgtaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg 5401 cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag 5461 gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcattt 5521 tagcttcctt agctcctgaa aatctcgata actcaaaaaa tacgcccggt agtgatctta 5581 tttcattatg gtgaaagttg gaacctctta cgtgccgatc aagtcaaaag cctccggtcg 5641 gaggcttttg actttctgct atggaggtca ggtatgattt aaatggtcag ttaga //