LOCUS Exported 5097 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Yeast Expression plasmid for SARS-CoV-2 nsp7. ACCESSION . VERSION . KEYWORDS pGBW-m4046264 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5097) TITLE SARS-CoV-2 protein Yeast expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5097) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5097 /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 724..741 /codon_start=1 /product="thrombin recognition and cleavage site" /label=thrombin site /translation="LVPRGS" CDS 742..807 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" primer_bind complement(852..870) /label=CYC1 /note="CYC1 transcription termination signal, reverse primer" CDS 1221..2024 /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 2107..3089 /label=2u ori /note="yeast 2u plasmid origin of replication" terminator 3167..3196 /label=T3Te terminator /note="phage T3 early transcription terminator" rep_origin 3218..3805 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" terminator 3817..3844 /label=T7Te terminator /note="phage T7 early transcription terminator" CDS complement(3868..4059) /codon_start=1 /gene="rop" /product="Rop protein, which maintains plasmids at low copy number" /label=rop /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA DELYRSCLARFGDDGENL" CDS complement(4060..4920) /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 4683..4702 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(4921..5023) /label=cat promoter /note="promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase" terminator 5024..5055 /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 aaaatgtcag atgtaaagtg cacatcagta gtcttactct cagttttgca acaactcaga 541 gtagaatcat catctaaatt gtgggctcaa tgtgtccagt tacacaatga cattctctta 601 gctaaagata ctactgaagc ctttgaaaaa atggtttcac tactttctgt tttgctttcc 661 atgcagggtg ctgtagacat aaacaagctt tgtgaagaaa tgctggacaa cagggcaacc 721 ttactcgtgc cgagaggttc tgattataaa gatcatgatg gtgattataa agatcatgat 781 attgattata aagatgatga tgataaataa acaggcccct tttcctttgt cgatatcatg 841 taattagtta tgtcacgctt acattcacgc cctcctccca catccgctct aaccgaaaag 901 gaaggagtta gacaacctga agtctaggtc cctatttatt ttttttaata gttatgttag 961 tattaagaac gttatttata tttcaaattt ttcttttttt tctgtacaaa gcggccgccg 1021 aaccactagt gcactgcagt acacggtttc cttgaaattt ttttgattcg gtaatctccg 1081 aacagaagga agaacgaagg aaggagcaca gacttagatt ggtatatata cgcatatgta 1141 gtgttgaaga aacatgaaat tgcccagtat tcttaaccca actgcacaga acaaaaacgt 1201 gcaggaaacg aagataaatc atgtcgaaag ctacatataa ggaacgtgct gctactcatc 1261 ctagtcctgt tgctgccaag ctatttaata tcatgcacga aaagcaaaca aacttgtgtg 1321 cttcattgga tgttcgtacc accaaggaat tactggagtt agttgaagca ttaggtccca 1381 aaatttgttt actaaaaaca catgtggata tcttgactga tttttccatg gagggcacag 1441 ttaagccgct aaaggcatta tccgccaagt acaatttttt actcttcgag gacagaaaat 1501 ttgctgacat tggtaataca gtcaaattgc agtactctgc gggtgtatac agaatagcag 1561 aatgggcaga cattacgaat gcacacggtg tggtgggccc aggtattgtt agcggtttga 1621 agcaggcggc agaagaagta acaaaggaac ctagaggcct tttgatgtta gcagaattgt 1681 catgcaaggg ctccctatct actggagaat atactaaggg tactgttgac attgcgaaga 1741 gcgacaaaga ttttgttatc ggctttattg ctcaaagaga catgggtgga agagatgaag 1801 gttacgattg gttgattatg acacccggtg tgggtttaga tgacaaggga gatgcattgg 1861 gtcaacagta tagaaccgtg gatgatgtgg tttctacagg atctgacatt attattgttg 1921 gaagaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt tacagaaaag 1981 caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact gtattataag 2041 taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat cagttattac 2101 ccgagtaacg aagcatctgt gcttcatttt gtagaacaaa aatgcaacgc gagagcgcta 2161 atttttcaaa caaagaatct gagctgcatt tttacagaac agaaatgcaa cgcgaaagcg 2221 ctattttacc aacgaagaat ctgtgcttca tttttgtaaa acaaaaatgc aacgcgagag 2281 cgctaatttt tcaaacaaag aatctgagct gcatttttac agaacagaaa tgcaacgcga 2341 gagcgctatt ttaccaacaa agaatctata cttctttttt gttctacaaa aatgcatccc 2401 gagagcgcta tttttctaac aaagcatctt agattacttt ttttctcctt tgtgcgctct 2461 ataatgcagt ctcttgataa ctttttgcac tgtaggtccg ttaaggttag aagaaggcta 2521 ctttggtgtc tattttctct tccataaaaa aagcctgact ccacttcccg cgtttactga 2581 ttactagcga agctgcgggt gcattttttc aagataaagg catccccgat tatattctat 2641 accgatgtgg attgcgcata ctttgtgaac agaaagtgat agcgttgatg attcttcatt 2701 ggtcagaaaa ttatgaacgg tttcttctat tttgtctcta tatactacgt ataggaaatg 2761 tttacatttt cgtattgttt tcgattcact ctatgaatag ttcttactac aatttttttg 2821 tctaaagagt aatactagag ataaacataa aaaatgtaga ggtcgagttt agatgcaagt 2881 tcaaggagcg aaaggtggat gggtaggtta tatagggata tagcacagag atatatagca 2941 aagagatact tttgagcaat gtttgtggaa gcggtattcg caatatttta gtagctcgtt 3001 acagtccggt gcgtttttgg ttttttgaaa gtgcgtcatc agagcgcttt tggttttcaa 3061 aagcgctctg aagttcctat actttctagc tagagaatag gaacttcccg agcggccgcg 3121 tgttacaagc ttgactccag cgtaactgga ctgcaatcaa ctcactggct caccttcacg 3181 ggtgggcctt tcttcggtag aaaatcaaag gatcttcttg agatcctttt tttctgcgcg 3241 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc 3301 aagagctacc aactcttttt ccgaggtaac tggcttcagc agagcgcaga taccaaatac 3361 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac 3421 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct 3481 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg 3541 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca 3601 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt 3661 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccaggggtaa acgcctggta 3721 tctttatagt cctgtcgggt ttcgccacct ctgacttgag catcgatttt tgtgatgctc 3781 gtcagggggg cggagcctat ggaaaaacgc cagcaacgca gaaaggccca cccgaaggtg 3841 agccaggtga ttacatttgg gccctcatca gaggttttca ccgtcatcac cgaaacgcgc 3901 gaggcagctg cggtaaagct catcagcgtg gtcgtgaagc gattcacaga tgtctgcctg 3961 ttcatccgcg tccagctcgt tgagtttctc cagaagcgtt aatgtctggc ttctgataaa 4021 gcgggccatg ttaagggcgg ttttttcctg tttggtcact taccaatgct taatcagtga 4081 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt 4141 gtagataact acgatgcggg agggcttacc atctggcccc agtgctgcaa tgataccgcg 4201 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg ggagggccga 4261 gcgcagaagt gatcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga 4321 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg 4381 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc 4441 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc 4501 gatcgttgcc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca 4561 taattctctt actgtcatgc catccgtgag atgcttttct gtgactggtg agtactcaac 4621 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg 4681 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc 4741 ggggcgtaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg 4801 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac 4861 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat 4921 tttagcttcc ttagctcctg aaaatctcga taactcaaaa aatacgcccg gtagtgatct 4981 tatttcatta tggtgaaagt tggaacctct tacgtgccga tcaagtcaaa agcctccggt 5041 cggaggcttt tgactttctg ctatggaggt caggtatgat ttaaatggtc agttaga //