LOCUS Exported 8514 bp ds-DNA circular SYN 13-SEP-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp2. ACCESSION . VERSION . KEYWORDS pGBW-m4133966 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8514) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 8514) AUTHORS . TITLE Direct Submission JOURNAL Exported Sep 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8514 /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 2494..2514 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 2515..2556 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" polyA_signal 2625..2746 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2636..2655) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2690..2709 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2861..2907 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2937..3073 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3118..3189 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(3415..4003) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3495..3514) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 4503..5712 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 5549..5568 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 5716..6744 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /label=HygR /note="confers resistance to hygromycin" /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA KE" polyA_signal 6771..6819 /note="synthetic polyadenylation signal" CDS 7085..7945 /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(7303..7322) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 7958..8094 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 8246..8317 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 8322..8368 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 8386..8434 /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 ccatgtacac tcgctatgtc 601 gataacaact tctgtggccc tgatggctac cctcttgagt gcattaaaga ccttctagca 661 cgtgctggta aagcttcatg cactttgtcc gaacaactgg actttattga cactaagagg 721 ggtgtatact gctgccgtga acatgagcat gaaattgctt ggtacacgga acgttctgaa 781 aagagctatg aattgcagac accttttgaa attaaattgg caaagaaatt tgacaccttc 841 aatggggaat gtccaaattt tgtatttccc ttaaattcca taatcaagac tattcaacca 901 agggttgaaa agaaaaagct tgatggcttt atgggtagaa ttcgatctgt ctatccagtt 961 gcgtcaccaa atgaatgcaa ccaaatgtgc ctttcaactc tcatgaagtg tgatcattgt 1021 ggtgaaactt catggcagac gggcgatttt gttaaagcca cttgcgaatt ttgtggcact 1081 gagaatttga ctaaagaagg tgccactact tgtggttact taccccaaaa tgctgttgtt 1141 aaaatttatt gtccagcatg tcacaattca gaagtaggac ctgagcatag tcttgccgaa 1201 taccataatg aatctggctt gaaaaccatt cttcgtaagg gtggtcgcac tattgccttt 1261 ggaggctgtg tgttctctta tgttggttgc cataacaagt gtgcctattg ggttccacgt 1321 gctagcgcta acataggttg taaccataca ggtgttgttg gagaaggttc cgaaggtctt 1381 aatgacaacc ttcttgaaat actccaaaaa gagaaagtca acatcaatat tgttggtgac 1441 tttaaactta atgaagagat cgccattatt ttggcatctt tttctgcttc cacaagtgct 1501 tttgtggaaa ctgtgaaagg tttggattat aaagcattca aacaaattgt tgaatcctgt 1561 ggtaatttta aagttacaaa aggaaaagct aaaaaaggtg cctggaatat tggtgaacag 1621 aaatcaatac tgagtcctct ttatgcattt gcatcagagg ctgctcgtgt tgtacgatca 1681 attttctccc gcactcttga aactgctcaa aattctgtgc gtgttttaca gaaggccgct 1741 ataacaatac tagatggaat ttcacagtat tcactgagac tcattgatgc tatgatgttc 1801 acatctgatt tggctactaa caatctagtt gtaatggcct acattacagg tggtgttgtt 1861 cagttgactt cgcagtggct aactaacatc tttggcactg tttatgaaaa actcaaaccc 1921 gtccttgatt ggcttgaaga gaagtttaag gaaggtgtag agtttcttag agacggttgg 1981 gaaattgtta aatttatctc aacctgtgct tgtgaaattg tcggtggaca aattgtcacc 2041 tgtgcaaagg aaattaagga gagtgttcag acattcttta agcttgtaaa taaatttttg 2101 gctttgtgtg ctgactctat cattattggt ggagctaaac ttaaagcctt gaatttaggt 2161 gaaacatttg tcacgcactc aaagggattg tacagaaagt gtgttaaatc cagagaagaa 2221 actggcctac tcatgcctct aaaagcccca aaagaaatta tcttcttaga gggagaaaca 2281 cttcccacag aagtgttaac agaggaagtt gtcttgaaaa ctggtgattt acaaccatta 2341 gaacaaccta ctagtgaagc tgttgaagct ccattggttg gtacaccagt ttgtattaac 2401 gggcttatgt tgctcgaaat caaagacaca gaaaagtact gtgcccttgc acctaatatg 2461 atggtaacaa acaatacctt cacactcaaa ggcgaaaacc tttactttca gtcggggaaa 2521 cctataccca atccacttct cggcttagat tctacctaag gatccgcagg cctctgctag 2581 cttgactgac tgagatacag cgtaccttca gctcacagac atgataagat acattgatga 2641 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga 2701 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg 2761 cattcatttt atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa 2821 cctctacaaa tgtggtattg gcccatctct atcggtatcg tagcataacc ccttggggcc 2881 tctaaacggg tcttgagggg ttttttgtgc ccctcgggcc ggattgctat ctaccggcat 2941 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga 3001 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga 3061 aatttatcct taaggtcgtc agctatcctg caggcgatct ctcgatttcg atcaagacat 3121 tcctttaatg gtcttttctg gacaccacta ggggtcagaa gtagttcatc aaactttctt 3181 ccctccctaa tctcattggt taccttgggc tatcgaaact taattaacca gtcaagtcag 3241 ctacttggcg agatcgactt gtctgggttt cgactacgct cagaattgcg tcagtcaagt 3301 tcgatctggt ccttgctatt gcacccgttc tccgattacg agtttcattt aaatcatgtg 3361 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca 3421 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa 3481 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc 3541 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc 3601 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct 3661 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg 3721 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag 3781 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta 3841 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg 3901 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt 3961 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt 4021 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag 4081 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat 4141 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc 4201 tatctcagcg atctgtctat ttcgttcatc catagttgca tttaaatttc cgaactctcc 4261 aaggccctcg tcggaaaatc ttcaaacctt tcgtccgatc catcttgcag gctacctctc 4321 gaacgaacta tcgcaagtct cttggccggc cttgcgcctt ggctattgct tggcagcgcc 4381 tatcgccagg tattactcca atcccgaata tccgagatcg ggatcacccg agagaagttc 4441 aacctacatc ctcaatcccg atctatccga gatccgagga atatcgaaat cggggcgcgc 4501 ctggcctccg cgccgggttt tggcgcctcc cgcgggcgcc cccctcgtca cggcgagcgc 4561 tgccacgtca gacgaagggc gcaggagcgt cctgatcctt ccgcccggac gctcaggaca 4621 gcggcccgct gctcataaga ctcggcctta gaaccccagt atcagcagaa ggacatttta 4681 ggacgggact tgggtgactc tagggcactg gttttctttc cagagagcgg aacaggcgag 4741 gaaaagtagt cccttctcgg cgattctgcg gagggatctc cgtggggcgg tgaacgccga 4801 tgattatata aggacgcgcc gggtgtggca cagctagttc cgtcgcagcc gggatttggg 4861 tcgcggttct tgtttgtgga tcgctgtgat cgtcacttgg tgagtagcgg gctgctgggc 4921 tggccggggc tttcgtggcc gccgggccgc tcggtgggac ggaagcgtgt ggagagaccg 4981 ccaagggctg tagtctgggt ccgcgagcaa ggttgccctg aactgggggt tggggggagc 5041 gcagcaaaat ggcggctgtt cccgagtctt gaatggaaga cgcttgtgag gcgggctgtg 5101 aggtcgttga aacaaggtgg ggggcatggt gggcggcaag aacccaaggt cttgagccct 5161 tcgctaatgc gggaaagctc ttattcgggt gagatgggct gggcaccatc tggggaccct 5221 gacgtgaagt ttgtcactga ctggagaact cggtttgtcg tctgttgcgg gggcggcagt 5281 tatggcggtg ccgttgggca gtgcacccgt acctttggga gcgcgcgccc tcgtcgtgtc 5341 gtgacgtcac ccgttctgtt ggcttataat gcagggtggg gccacctgcc ggtaggtgtg 5401 cggtaggctt ttctccgtcg caggacgcag ggttcgggcc tagggtaggc tctcctgaat 5461 cgacaggcgc cggacctctg gtgaggggag ggataagtga ggcgtcagtt tctttggtcg 5521 gttttatgta cctatcttct taagtagctg aagctccggt tttgaactat gcgctcgggg 5581 ttggcgagtg tgttttgtga agttttttag gcaccttttg aaatgtaatc atttgggtca 5641 atatgtaatt ttcagtgtta gacttgtaaa ttgtccgcta aattctggcc gtttttggct 5701 tttttgttag acaacatggg taaaaagcct gaactcaccg cgacgtctgt cgagaagttt 5761 ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc tctcggaggg cgaagaatct 5821 cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc tgcgggtaaa tagctgcgcc 5881 gatggtttct acaaagatcg ttatgtttat cggcactttg catccgccgc gctcccgatt 5941 ccggaagtgc ttgacattgg ggagttcagc gagagcctga cctattgcat ctcccgccgt 6001 gcacagggtg tcacgttgca agacctgcct gaaaccgaac tgcccgctgt tctgcagccg 6061 gtcgcggagg caatggatgc catcgctgcc gccgatctta gccagacgag cgggttcggc 6121 ccattcggac cgcaaggaat cggtcaatac actacatggc gtgatttcat atgcgcgatt 6181 gctgatcccc atgtgtatca ctggcaaact gtgatggacg acaccgtcag tgcgtccgtc 6241 gcgcaggctc tcgatgagct gatgctttgg gccgaggact gccccgaagt ccggcacctc 6301 gtgcacgcgg atttcggctc caacaatgtc ctgacggaca atggccgcat aacagcggtc 6361 attgactgga gcgaggcgat gttcggggat tcccaatacg aggtcgccaa catcttcttc 6421 tggaggccgt ggttggcttg tatggagcag cagacgcgct acttcgagcg gaggcatccg 6481 gagcttgcag gatcgccccg gctccgggcg tatatgctcc gcattggtct tgaccaactc 6541 tatcagagct tggttgacgg caatttcgat gatgcagctt gggcgcaggg tcgatgcgac 6601 gcaatcgtcc gatccggagc cgggactgtc gggcgtacac aaatcgcccg cagaagcgcc 6661 gccgtctgga ccgatggctg tgtagaagta ctcgccgata gtggaaaccg acgccccagc 6721 actcgtccga gggcaaagga ataagctagt atgtaagcct agtcttagat aataaaatcg 6781 ctatccatcg aagatggatg tgtgttggtt ttttgtgtgt gtaacgctag gcgcgcctgg 6841 tgtaccgaga acgatcctct cagtgcgagt ctcgacgatc catatcgttg cttggcagtc 6901 agccagtcgg aatccagctt gggacccagg aagtccaatc gtcagatatt gtactcaagc 6961 ctggtcacgg cagcgtaccg atctgtttaa acctagatat tgatagtctg atcggtcaac 7021 gtataatcga gtcctagctt ttgcaaacat ctatcaagag acaggatcag caggaggctt 7081 tcgcatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct 7141 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg 7201 tgcgcgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg 7261 ccccgaagaa cgctttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt 7321 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga 7381 cttggttgag tattcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga 7441 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac 7501 gattggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg 7561 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac 7621 gatgcctgta gcaatggcaa caaccttgcg taaactatta actggcgaac tacttactct 7681 agcttcccgg caacagttga tagactggat ggaggcggat aaagttgcag gaccacttct 7741 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg 7801 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat 7861 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg 7921 tgcctcactg attaagcatt ggtaaccgat tctaggtgca ttggcgcaga aaaaaatgcc 7981 tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac ggatacggct 8041 tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc ttaagatccc 8101 gaatcgttta aactcgactc tggctctatc gaatctccgt cgtttcgagc ttacgcgaac 8161 agccgtggcg ctcatttgct cgtcgggcat cgaatctcgt cagctatcgt cagcttacct 8221 ttttggcagc gatcgcggct cccgacatct tggaccatta gctccacagg tatcttcttc 8281 cctctagtgg tcataacagc agcttcagct acctctcaat tcaaaaaacc cctcaagacc 8341 cgtttagagg ccccaagggg ttatgctatc aatcgttgcg ttacacacac aaaaaaccaa 8401 cacacatcca tcttcgatgg atagcgattt tattatctaa ctgctgatcg agtgtagcca 8461 gatctagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccg //