LOCUS Exported 6144 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp2. ACCESSION . VERSION . KEYWORDS pGBW-m4134161 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6144) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6144) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6144 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 1922..1942 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1943..1960 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 2013..2134 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2024..2043) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2078..2097 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2249..2295 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2325..2461 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2506..2577 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2803..3391) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2883..2902) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 4137..4997 /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(4355..4374) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5010..5146 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 5298..5369 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 5374..5420 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5438..5486 /note="synthetic polyadenylation signal" enhancer 5567..5870 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 5872..6074 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6025..6045 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 6143..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggcc tacacaagat atgtggataa caatttttgt gggccggacg gatacccttt 61 agaatgcatc aaagatttgc ttgcaagggc gggcaaggct tcgtgcactc tgtccgagca 121 gctggacttc attgatacca agcgcggtgt ttattgttgc cgagagcatg aacacgagat 181 agcctggtat acggaacggt cagagaaaag ttacgaactc caaaccccat tcgagatcaa 241 gctagctaaa aagtttgaca ctttcaacgg cgaatgtccc aattttgtct ttcccctgaa 301 ctctattatc aaaacaatac agcctcgggt ggagaagaag aaattagacg gattcatggg 361 gcgtattagg agcgtatacc cagtggccag cccgaatgaa tgcaaccaga tgtgtctttc 421 cacactgatg aagtgcgatc actgtggcga gacttcttgg caaaccggtg acttcgtaaa 481 agcgacatgc gagttttgtg gtaccgagaa tctgactaag gagggagcta ccacatgcgg 541 ctatttgcca cagaatgcag tggtcaagat ctactgtcct gcatgccata actctgaagt 601 cggccccgag cactcgctcg ccgaatatca caatgagagc gggctaaaaa ctatactgag 661 gaaaggaggt cgcaccatcg cctttggggg ctgcgttttc tcctacgtgg ggtgtcataa 721 caaatgtgcc tattgggttc ccagagcctc cgctaacatt gggtgcaatc atacgggagt 781 ggtcggggaa ggctcagaag gcttgaacga taacctcctt gaaattctgc aaaaggagaa 841 ggtgaatatc aatatcgttg gcgacttcaa gttaaatgag gaaatcgcga ttattctggc 901 aagtttttct gcctcaacta gcgcctttgt agagacagtg aaaggactcg attataaagc 961 tttcaaacag atagtcgaga gttgtggtaa cttcaaggtt acaaagggca aggctaaaaa 1021 gggtgcttgg aacattggag agcagaagtc aatcttgagt ccactgtacg cgtttgcatc 1081 tgaggccgcc cgtgtggtca ggagcatatt cagtcgaacc ctagaaaccg cacagaatag 1141 cgtgcgggta ttacagaagg cggctattac tattctcgac ggtatctcgc agtattctct 1201 gagactgatc gacgccatga tgtttacaag cgatctggca acgaataact tagtggtcat 1261 ggcgtacata accggagggg tcgtgcaact tacctcccag tggctaacga acatctttgg 1321 gacggtttac gagaagctaa aacctgtgct ggactggctc gaagagaaat tcaaagaagg 1381 cgtggaattt ttgcgtgacg ggtgggagat cgtaaagttc atatcaacat gtgcttgcga 1441 aatcgttggc ggacaaatcg taacttgcgc aaaagagatt aaagaatcgg tgcagacctt 1501 ctttaaactt gttaacaaat tcctggcttt gtgtgccgat tctattatca tcgggggagc 1561 aaagcttaaa gccctaaatt taggcgagac atttgtgacg cacagtaagg gcctgtatag 1621 aaagtgcgtg aagagtcgcg aagaaaccgg actgctcatg cccttaaaag cgccaaaaga 1681 aataattttc ctggaagggg agacattgcc cactgaagtg ctcactgagg aagtagttct 1741 gaaaactggg gatctgcaac ctttagagca gccgacctcc gaggccgtcg aggcacctct 1801 cgtgggaacc ccggtctgta ttaatgggtt gatgctgctc gagattaagg ataccgaaaa 1861 atactgcgca ctagccccca acatgatggt gactaacaac acatttaccc tcaagggagg 1921 ggaaaacctt tactttcagt cgcaccatca ccatcaccat taagctagct tgactgactg 1981 agatacagcg taccttcagc tcacagacat gataagatac attgatgagt ttggacaaac 2041 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt 2101 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat 2161 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg 2221 tggtattggc ccatctctat cggtatcgta gcataacccc ttggggcctc taaacgggtc 2281 ttgaggggtt ttttgtgccc ctcgggccgg attgctatct accggcattg gcgcagaaaa 2341 aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga 2401 tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta 2461 aggtcgtcag ctatcctgca ggcgatctct cgatttcgat caagacattc ctttaatggt 2521 cttttctgga caccactagg ggtcagaagt agttcatcaa actttcttcc ctccctaatc 2581 tcattggtta ccttgggcta tcgaaactta attaaccagt caagtcagct acttggcgag 2641 atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc gatctggtcc 2701 ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag caaaaggcca 2761 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc 2821 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact 2881 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct 2941 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag 3001 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca 3061 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa 3121 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc 3181 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag 3241 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg 3301 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca 3361 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc 3421 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag 3481 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata 3541 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat 3601 ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa ggccctcgtc 3661 ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga acgaactatc 3721 gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta tcgccaggta 3781 ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa cctacatcct 3841 caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct ggtgtaccga 3901 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc 3961 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac 4021 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc 4081 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga 4141 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt 4201 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag 4261 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag 4321 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta 4381 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg 4441 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca 4501 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag 4561 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc 4621 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg 4681 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc 4741 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg 4801 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg 4861 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga 4921 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac 4981 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga 5041 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac 5101 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc ccgaatcgtt 5161 taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg 5221 cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac ctttttggca 5281 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt 5341 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga 5401 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc 5461 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctagt 5521 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta 5581 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga 5641 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt 5701 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta 5761 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg 5821 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg ctgatgcggt 5881 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc 5941 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat 6001 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct 6061 atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca 6121 ctcgacacac ccgccagcgg ccgc //