LOCUS Exported 6270 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Yeast expression plasmid for SARS-CoV-2 nsp4. ACCESSION . VERSION . KEYWORDS pGBW-m4046697 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6270) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6270) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6270 /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" primer_bind complement(2025..2043) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 2394..3197 /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 3280..4262 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 4340..4369 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 4391..4978 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 4990..5017 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(5041..5232) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(5233..6093) /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 5856..5875 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(6094..6196) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 6197..6228 /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 aagattgtca acaattggct taaacagttg ataaaggtta ctttagtatt tttgttcgtt 541 gcagccatct tctatctaat tacacctgtc cacgtgatga gtaagcatac cgacttttcc 601 tcagagatca ttggttacaa agctatcgat ggcggagtta ctagagatat tgcttcgacg 661 gacacctgtt tcgctaacaa gcacgcggac tttgatactt ggttctctca aaggggtggt 721 tcctacacaa acgacaaggc ctgccccttg attgcagctg ttatcactag agaagttggt 781 ttcgtcgtgc caggtttacc agggactata ctgcgtacca ccaatggtga ttttttgcat 841 ttcttaccaa gagtcttctc tgctgttggc aatatttgtt atacccctag caaattgatc 901 gaatacactg attttgccac ctctgcttgt gtattggctg ctgaatgcac tattttcaag 961 gatgcaagtg gtaagccagt tccatactgt tatgacacaa acgtcctcga aggttctgtt 1021 gcttacgaat ccttgcgacc ggacactaga tatgtgttaa tggatggatc aatcattcaa 1081 tttccaaaca cataccttga aggttctgtt cgggttgtca ccaccttcga ctccgagtac 1141 tgtcgccacg gtacttgtga aagatcagaa gccggtgtct gtgtttctac cagcggtaga 1201 tgggtgttga ataacgatta ttaccgttct cttccaggtg tcttctgcgg tgtcgatgct 1261 gttaatctat tgactaacat gtttacccct ttgatccaac caataggtgc gctggacatt 1321 tccgcatcta ttgtcgccgg tggaatcgtt gctatcgttg ttacttgttt ggcttactac 1381 ttcatgagat ttagaagagc ctttggggag tattcgcatg tcgtggcttt caacactcta 1441 ttatttttga tgtcgttcac tgttttgtgt cttacaccag tatactcttt cttgcctggt 1501 gtatactccg ttatctattt atacctgaca ttctacctga ctaacgatgt cagctttttg 1561 gcacacattc aatggatggt aatgttcacc ccattggttc ccttctggat aacgatcgct 1621 tacatcattt gcataagtac taaacatttc tattggtttt tctcaaatta tctaaagaga 1681 agagttgtct ttaatggtgt cagcttttct accttcgaag aagctgcctt gtgtaccttt 1741 ttgctcaaca aagaaatgta cttgaagttg agatctgacg ttttactccc gctaacccaa 1801 tataacagat atttagcttt atacaacaag tacaaatact tctcgggtgc catggatact 1861 acttcatacc gtgaagcggc ttgttgtcac cttgctaagg ctttgaatga tttctctaat 1921 tccggttccg acgtcttgta tcaaccacct cagacgtcta ttacctccgc tgttttgcaa 1981 taaacaggcc ccttttcctt tgtcgatatc atgtaattag ttatgtcacg cttacattca 2041 cgccctcctc ccacatccgc tctaaccgaa aaggaaggag ttagacaacc tgaagtctag 2101 gtccctattt atttttttta atagttatgt tagtattaag aacgttattt atatttcaaa 2161 tttttctttt ttttctgtac aaagcggccg ccgaaccact agtgcactgc agtacacggt 2221 ttccttgaaa tttttttgat tcggtaatct ccgaacagaa ggaagaacga aggaaggagc 2281 acagacttag attggtatat atacgcatat gtagtgttga agaaacatga aattgcccag 2341 tattcttaac ccaactgcac agaacaaaaa cgtgcaggaa acgaagataa atcatgtcga 2401 aagctacata taaggaacgt gctgctactc atcctagtcc tgttgctgcc aagctattta 2461 atatcatgca cgaaaagcaa acaaacttgt gtgcttcatt ggatgttcgt accaccaagg 2521 aattactgga gttagttgaa gcattaggtc ccaaaatttg tttactaaaa acacatgtgg 2581 atatcttgac tgatttttcc atggagggca cagttaagcc gctaaaggca ttatccgcca 2641 agtacaattt tttactcttc gaggacagaa aatttgctga cattggtaat acagtcaaat 2701 tgcagtactc tgcgggtgta tacagaatag cagaatgggc agacattacg aatgcacacg 2761 gtgtggtggg cccaggtatt gttagcggtt tgaagcaggc ggcagaagaa gtaacaaagg 2821 aacctagagg ccttttgatg ttagcagaat tgtcatgcaa gggctcccta tctactggag 2881 aatatactaa gggtactgtt gacattgcga agagcgacaa agattttgtt atcggcttta 2941 ttgctcaaag agacatgggt ggaagagatg aaggttacga ttggttgatt atgacacccg 3001 gtgtgggttt agatgacaag ggagatgcat tgggtcaaca gtatagaacc gtggatgatg 3061 tggtttctac aggatctgac attattattg ttggaagagg actatttgca aagggaaggg 3121 atgctaaggt agagggtgaa cgttacagaa aagcaggctg ggaagcatat ttgagaagat 3181 gcggccagca aaactaaaaa actgtattat aagtaaatgc atgtatacta aactcacaaa 3241 ttagagcttc aatttaatta tatcagttat tacccgagta acgaagcatc tgtgcttcat 3301 tttgtagaac aaaaatgcaa cgcgagagcg ctaatttttc aaacaaagaa tctgagctgc 3361 atttttacag aacagaaatg caacgcgaaa gcgctatttt accaacgaag aatctgtgct 3421 tcatttttgt aaaacaaaaa tgcaacgcga gagcgctaat ttttcaaaca aagaatctga 3481 gctgcatttt tacagaacag aaatgcaacg cgagagcgct attttaccaa caaagaatct 3541 atacttcttt tttgttctac aaaaatgcat cccgagagcg ctatttttct aacaaagcat 3601 cttagattac tttttttctc ctttgtgcgc tctataatgc agtctcttga taactttttg 3661 cactgtaggt ccgttaaggt tagaagaagg ctactttggt gtctattttc tcttccataa 3721 aaaaagcctg actccacttc ccgcgtttac tgattactag cgaagctgcg ggtgcatttt 3781 ttcaagataa aggcatcccc gattatattc tataccgatg tggattgcgc atactttgtg 3841 aacagaaagt gatagcgttg atgattcttc attggtcaga aaattatgaa cggtttcttc 3901 tattttgtct ctatatacta cgtataggaa atgtttacat tttcgtattg ttttcgattc 3961 actctatgaa tagttcttac tacaattttt ttgtctaaag agtaatacta gagataaaca 4021 taaaaaatgt agaggtcgag tttagatgca agttcaagga gcgaaaggtg gatgggtagg 4081 ttatataggg atatagcaca gagatatata gcaaagagat acttttgagc aatgtttgtg 4141 gaagcggtat tcgcaatatt ttagtagctc gttacagtcc ggtgcgtttt tggttttttg 4201 aaagtgcgtc atcagagcgc ttttggtttt caaaagcgct ctgaagttcc tatactttct 4261 agctagagaa taggaacttc ccgagcggcc gcgtgttaca agcttgactc cagcgtaact 4321 ggactgcaat caactcactg gctcaccttc acgggtgggc ctttcttcgg tagaaaatca 4381 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac 4441 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaggt 4501 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg 4561 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc 4621 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt 4681 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga 4741 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct 4801 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg 4861 cacgagggag cttccagggg taaacgcctg gtatctttat agtcctgtcg ggtttcgcca 4921 cctctgactt gagcatcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa 4981 cgccagcaac gcagaaaggc ccacccgaag gtgagccagg tgattacatt tgggccctca 5041 tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc 5101 gtggtcgtga agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt 5161 ctccagaagc gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc 5221 ctgtttggtc acttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt 5281 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatgc gggagggctt 5341 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt 5401 atcagcaata aaccagccag ccgggagggc cgagcgcaga agtgatcctg caactttatc 5461 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa 5521 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg 5581 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt 5641 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gccagaagta agttggccgc 5701 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt 5761 gagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg 5821 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac 5881 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcgt aaactctcaa ggatcttacc 5941 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt 6001 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg 6061 aataagggcg acacggaaat gttgaatact cattttagct tccttagctc ctgaaaatct 6121 cgataactca aaaaatacgc ccggtagtga tcttatttca ttatggtgaa agttggaacc 6181 tcttacgtgc cgatcaagtc aaaagcctcc ggtcggaggc ttttgacttt ctgctatgga 6241 ggtcaggtat gatttaaatg gtcagttaga //