LOCUS Exported 4824 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp8. ACCESSION . VERSION . KEYWORDS pGBW-m4134147 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4824) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 4824) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4824 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 602..622 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 623..640 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 693..814 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(704..723) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 758..777 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 929..975 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1005..1141 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1186..1257 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1483..2071) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1563..1582) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 2817..3677 /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(3035..3054) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 3690..3826 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3978..4049 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4054..4100 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 4118..4166 /note="synthetic polyadenylation signal" enhancer 4247..4550 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4552..4754 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4705..4725 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 4823..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggca attgcctcag aatttagttc cctcccctcg tacgctgcgt tcgctaccgc 61 ccaagaggca tatgagcagg ccgtagccaa cggggacagc gaagtggttc ttaagaaact 121 gaagaaatct ttgaatgtgg caaagagcga gtttgatcgg gacgcggcaa tgcagaggaa 181 gctggaaaaa atggctgatc aggctatgac acagatgtac aaacaagccc gatccgagga 241 caagagagct aaagtcactt ctgccatgca aacgatgcta ttcaccatgt tacgtaagct 301 ggataacgac gcactcaata acataatcaa taacgcccgc gatggttgtg tgccattgaa 361 tatcattcct cttactacag cggccaagct gatggtcgtt atcccggact ataacacata 421 caaaaatacc tgcgacggca ccactttcac gtatgctagc gccctgtggg aaatacagca 481 ggtagtggat gccgattcaa agattgtgca gcttagtgag atctccatgg acaacagtcc 541 caacttggct tggcctttaa tcgtcacagc tctcagagcc aatagcgcag ttaaactgca 601 agaaaacctt tactttcagt cgcaccatca ccatcaccat taagctagct tgactgactg 661 agatacagcg taccttcagc tcacagacat gataagatac attgatgagt ttggacaaac 721 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt 781 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat 841 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg 901 tggtattggc ccatctctat cggtatcgta gcataacccc ttggggcctc taaacgggtc 961 ttgaggggtt ttttgtgccc ctcgggccgg attgctatct accggcattg gcgcagaaaa 1021 aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga 1081 tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta 1141 aggtcgtcag ctatcctgca ggcgatctct cgatttcgat caagacattc ctttaatggt 1201 cttttctgga caccactagg ggtcagaagt agttcatcaa actttcttcc ctccctaatc 1261 tcattggtta ccttgggcta tcgaaactta attaaccagt caagtcagct acttggcgag 1321 atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc gatctggtcc 1381 ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag caaaaggcca 1441 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc 1501 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact 1561 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct 1621 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag 1681 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca 1741 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa 1801 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc 1861 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag 1921 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg 1981 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca 2041 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc 2101 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag 2161 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata 2221 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat 2281 ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa ggccctcgtc 2341 ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga acgaactatc 2401 gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta tcgccaggta 2461 ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa cctacatcct 2521 caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct ggtgtaccga 2581 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc 2641 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac 2701 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc 2761 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga 2821 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt 2881 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag 2941 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag 3001 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta 3061 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg 3121 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca 3181 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag 3241 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc 3301 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg 3361 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc 3421 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg 3481 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg 3541 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga 3601 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac 3661 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga 3721 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac 3781 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc ccgaatcgtt 3841 taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg 3901 cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac ctttttggca 3961 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt 4021 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga 4081 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc 4141 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctagt 4201 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta 4261 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga 4321 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt 4381 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta 4441 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg 4501 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg ctgatgcggt 4561 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc 4621 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat 4681 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct 4741 atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca 4801 ctcgacacac ccgccagcgg ccgc //