LOCUS Exported 5199 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 3C-like proteinase. ACCESSION . VERSION . KEYWORDS pGBW-m4133872 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5199) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5199) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5199 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 926..946 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 950..1015 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" polyA_signal 1068..1189 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1079..1098) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1133..1152 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1304..1350 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1380..1516 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1561..1632 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1858..2446) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1938..1957) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 3192..4052 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(3410..3429) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 4065..4201 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 4353..4424 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4429..4475 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 4493..4541 /note="synthetic polyadenylation signal" enhancer 4622..4925 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4927..5129 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 5080..5100 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgagt gggttccgga aaatggcgtt tccctccggt aaggtcgaag gatgcatggt 61 tcaggtgacc tgtggcacaa ctacgttaaa cggcctatgg ctggacgatg tggtatactg 121 tcctcgacac gtcatctgca catcagagga catgctgaat ccaaactatg aggatttgct 181 cataagaaag agcaatcata actttctggt gcaagctgga aatgtacagc ttagggttat 241 tgggcactct atgcagaact gcgtgttaaa actcaaggtg gataccgcca atccgaaaac 301 acctaagtac aaattcgtgc gcatccagcc aggccaaacc ttctctgtcc tggcatgtta 361 taacggcagc ccctcaggtg tctaccagtg cgccatgcgc cctaatttta ctattaaggg 421 ttccttcttg aacgggtcgt gcggatcggt cggctttaat atcgactatg actgtgtgag 481 tttttgctac atgcatcaca tggaacttcc gactggggtt cacgctggaa cggatctgga 541 gggtaacttc tatggaccct tcgtggatag acagaccgcc caggcagccg gcactgacac 601 taccattacg gtgaatgttc tagcttggct ctacgcagct gtaataaacg gcgatcgttg 661 gttcttgaac aggtttacaa caaccctgaa cgactttaat ctagtcgcca tgaaatataa 721 ttatgaacca ctgacccaag atcatgtaga cattctcggg ccactgagcg cacagacagg 781 gatcgctgtg ctggacatgt gtgccagttt aaaggagctg cttcagaatg gtatgaacgg 841 ccgaacaatc ctagggtctg cgctcttaga ggatgaattt acccccttcg atgtagtgcg 901 gcaatgtagc ggcgttactt ttcaagaaaa cctttacttt cagtcgatgg attataagga 961 ccacgatgga gactacaaag atcatgacat tgattacaag gacgacgatg acaaataagc 1021 tagcttgact gactgagata cagcgtacct tcagctcaca gacatgataa gatacattga 1081 tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg 1141 tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa 1201 ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta 1261 aaacctctac aaatgtggta ttggcccatc tctatcggta tcgtagcata accccttggg 1321 gcctctaaac gggtcttgag gggttttttg tgcccctcgg gccggattgc tatctaccgg 1381 cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc cacatatgcc 1441 agattcagca acggatacgg cttccccaac ttgcccactt ccatacgtgt cctccttacc 1501 agaaatttat ccttaaggtc gtcagctatc ctgcaggcga tctctcgatt tcgatcaaga 1561 cattccttta atggtctttt ctggacacca ctaggggtca gaagtagttc atcaaacttt 1621 cttccctccc taatctcatt ggttaccttg ggctatcgaa acttaattaa ccagtcaagt 1681 cagctacttg gcgagatcga cttgtctggg tttcgactac gctcagaatt gcgtcagtca 1741 agttcgatct ggtccttgct attgcacccg ttctccgatt acgagtttca tttaaatcat 1801 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt 1861 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg 1921 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc 1981 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt 2041 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa 2101 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta 2161 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa 2221 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa 2281 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt 2341 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt 2401 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat 2461 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat 2521 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc 2581 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc 2641 acctatctca gcgatctgtc tatttcgttc atccatagtt gcatttaaat ttccgaactc 2701 tccaaggccc tcgtcggaaa atcttcaaac ctttcgtccg atccatcttg caggctacct 2761 ctcgaacgaa ctatcgcaag tctcttggcc ggccttgcgc cttggctatt gcttggcagc 2821 gcctatcgcc aggtattact ccaatcccga atatccgaga tcgggatcac ccgagagaag 2881 ttcaacctac atcctcaatc ccgatctatc cgagatccga ggaatatcga aatcggggcg 2941 cgcctggtgt accgagaacg atcctctcag tgcgagtctc gacgatccat atcgttgctt 3001 ggcagtcagc cagtcggaat ccagcttggg acccaggaag tccaatcgtc agatattgta 3061 ctcaagcctg gtcacggcag cgtaccgatc tgtttaaacc tagatattga tagtctgatc 3121 ggtcaacgta taatcgagtc ctagcttttg caaacatcta tcaagagaca ggatcagcag 3181 gaggctttcg catgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat 3241 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc 3301 agttgggtgc gcgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga 3361 gttttcgccc cgaagaacgc tttccaatga tgagcacttt taaagttctg ctatgtggcg 3421 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc 3481 agaatgactt ggttgagtat tcaccagtca cagaaaagca tcttacggat ggcatgacag 3541 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc 3601 tgacaacgat tggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg 3661 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg 3721 acaccacgat gcctgtagca atggcaacaa ccttgcgtaa actattaact ggcgaactac 3781 ttactctagc ttcccggcaa cagttgatag actggatgga ggcggataaa gttgcaggac 3841 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg 3901 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg 3961 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg 4021 agataggtgc ctcactgatt aagcattggt aaccgattct aggtgcattg gcgcagaaaa 4081 aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga 4141 tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta 4201 agatcccgaa tcgtttaaac tcgactctgg ctctatcgaa tctccgtcgt ttcgagctta 4261 cgcgaacagc cgtggcgctc atttgctcgt cgggcatcga atctcgtcag ctatcgtcag 4321 cttacctttt tggcagcgat cgcggctccc gacatcttgg accattagct ccacaggtat 4381 cttcttccct ctagtggtca taacagcagc ttcagctacc tctcaattca aaaaacccct 4441 caagacccgt ttagaggccc caaggggtta tgctatcaat cgttgcgtta cacacacaaa 4501 aaaccaacac acatccatct tcgatggata gcgattttat tatctaactg ctgatcgagt 4561 gtagccagat ctagtaatca attacggggt cattagttca tagcccatat atggagttcc 4621 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat 4681 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc 4741 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc 4801 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt 4861 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta 4921 ccatgctgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg 4981 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac 5041 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg 5101 tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc agatctttgt 5161 cgatcctacc atccactcga cacacccgcc agcggccgc //