LOCUS Exported 5190 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp8. ACCESSION . VERSION . KEYWORDS pGBW-m4133314 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5190) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5190) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5190 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 1..304 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 306..508 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 459..479 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" CDS 1174..1194 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1543..1560 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 1637..1758 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1648..1667) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1702..1721 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1873..1919 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1949..2085 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2130..2201 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2427..3015) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2507..2526) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 3761..4621 /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(3979..3998) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 4634..4770 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 4922..4993 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4998..5044 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5062..5110 /note="synthetic polyadenylation signal" ORIGIN 1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 301 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc 541 gatcctacca tccactcgac acacccgcca gcggccgcca ccatgatagc ctcagagttt 601 agttcccttc catcatatgc agcttttgct actgctcaag aagcttatga gcaggctgtt 661 gctaatggtg attctgaagt tgttcttaaa aagttgaaga agtctttgaa tgtggctaaa 721 tctgaatttg accgtgatgc agccatgcaa cgtaagttgg aaaagatggc tgatcaagct 781 atgacccaaa tgtataaaca ggctagatct gaggacaaga gggcaaaagt tactagtgct 841 atgcagacaa tgcttttcac tatgcttaga aagttggata atgatgcact caacaacatt 901 atcaacaatg caagagatgg ttgtgttccc ttgaacataa tacctcttac aacagcagcc 961 aaactaatgg ttgtcatacc agactataac acatataaaa atacgtgtga tggtacaaca 1021 tttacttatg catcagcatt gtgggaaatc caacaggttg tagatgcaga tagtaaaatt 1081 gttcaactta gtgaaattag tatggacaat tcacctaatt tagcatggcc tcttattgta 1141 acagctttaa gggccaattc tgctgtcaaa ttagaaaacc tttactttca gtcgatgagt 1201 gacaaaatca ttcatctaac cgatgactcc tttgatacag acgtgttgaa ggccgatggt 1261 gcgatactgg tagatttctg ggctgaatgg tgtggcccgt gcaagatgat cgcacccatt 1321 ctggacgaga tcgccgacga gtatcaagga aaattaactg tcgctaagct taatatagat 1381 cagaacccag ggacggcacc taaatacggc atcaggggaa tacctaccct cctgctcttt 1441 aagaatgggg aagttgccgc gacaaaagtg ggagccctgt ctaagggcca gctaaaagag 1501 ttcttggacg ctaacctggc aggttcaggc tcggggcaca tgcaccatca ccatcaccat 1561 agcagcgggc tcgtgccacg cggctaagct agcttgactg actgagatac agcgtacctt 1621 cagctcacag acatgataag atacattgat gagtttggac aaaccacaac tagaatgcag 1681 tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt aaccattata 1741 agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca ggttcagggg 1801 gaggtgtggg aggtttttta aagcaagtaa aacctctaca aatgtggtat tggcccatct 1861 ctatcggtat cgtagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgt 1921 gcccctcggg ccggattgct atctaccggc attggcgcag aaaaaaatgc ctgatgcgac 1981 gctgcgcgtc ttatactccc acatatgcca gattcagcaa cggatacggc ttccccaact 2041 tgcccacttc catacgtgtc ctccttacca gaaatttatc cttaaggtcg tcagctatcc 2101 tgcaggcgat ctctcgattt cgatcaagac attcctttaa tggtcttttc tggacaccac 2161 taggggtcag aagtagttca tcaaactttc ttccctccct aatctcattg gttaccttgg 2221 gctatcgaaa cttaattaac cagtcaagtc agctacttgg cgagatcgac ttgtctgggt 2281 ttcgactacg ctcagaattg cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt 2341 tctccgatta cgagtttcat ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac 2401 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac 2461 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg 2521 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac 2581 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat 2641 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag 2701 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac 2761 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt 2821 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt 2881 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc 2941 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga 3001 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac 3061 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc 3121 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct 3181 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca 3241 tccatagttg catttaaatt tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc 3301 tttcgtccga tccatcttgc aggctacctc tcgaacgaac tatcgcaagt ctcttggccg 3361 gccttgcgcc ttggctattg cttggcagcg cctatcgcca ggtattactc caatcccgaa 3421 tatccgagat cgggatcacc cgagagaagt tcaacctaca tcctcaatcc cgatctatcc 3481 gagatccgag gaatatcgaa atcggggcgc gcctggtgta ccgagaacga tcctctcagt 3541 gcgagtctcg acgatccata tcgttgcttg gcagtcagcc agtcggaatc cagcttggga 3601 cccaggaagt ccaatcgtca gatattgtac tcaagcctgg tcacggcagc gtaccgatct 3661 gtttaaacct agatattgat agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc 3721 aaacatctat caagagacag gatcagcagg aggctttcgc atgagtattc aacatttccg 3781 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac 3841 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgcg cgagtgggtt acatcgaact 3901 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgct ttccaatgat 3961 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga 4021 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac 4081 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat 4141 gagtgataac actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac 4201 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct 4261 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac 4321 cttgcgtaaa ctattaactg gcgaactact tactctagct tcccggcaac agttgataga 4381 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg 4441 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact 4501 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac 4561 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta 4621 accgattcta ggtgcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata 4681 ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc acttccatac 4741 gtgtcctcct taccagaaat ttatccttaa gatcccgaat cgtttaaact cgactctggc 4801 tctatcgaat ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc 4861 gggcatcgaa tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg 4921 acatcttgga ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct 4981 tcagctacct ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat 5041 gctatcaatc gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag 5101 cgattttatt atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc 5161 attagttcat agcccatata tggagttccg //