LOCUS Exported 6485 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp9. ACCESSION . VERSION . KEYWORDS pGBW-m4134408 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6485) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6485) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6485 /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 925..945 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 946..972 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" polyA_signal 1025..1146 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1036..1055) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1090..1109 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1261..1307 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1337..1473 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1518..1589 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1815..2403) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1895..1914) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 2903..4112 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3949..3968 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 4116..4715 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(4116..4135) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4612..4632 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 4742..4790 /note="synthetic polyadenylation signal" CDS 5056..5916 /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(5274..5293) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5929..6065 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6217..6288 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6293..6339 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6357..6405 /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 ccatgaataa cgagttaagc 601 ccagtagcgc ttcgccagat gagttgcgcc gcaggtacaa cccaaactgc ttgtacggat 661 gacaacgccc tcgcttacta taataccaca aagggaggcc gattcgttct ggccctgttg 721 tcggatctgc aagacttgaa atgggcacgt tttcccaagt ccgacgggac tggcaccatt 781 tacacagaac tagagccgcc atgcaggttc gtgactgata cccctaaagg acccaaggtc 841 aaatatctgt attttatcaa ggggcttaac aatctcaata gaggcatggt gttaggatct 901 ctagccgcaa cggtgcggct gcaagaaaac ctttactttc agtcgtaccc ctatgacgta 961 ccagattatg cgtaagctag cttgactgac tgagatacag cgtaccttca gctcacagac 1021 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc 1081 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa 1141 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag 1201 gttttttaaa gcaagtaaaa cctctacaaa tgtggtattg gcccatctct atcggtatcg 1261 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgtgc ccctcgggcc 1321 ggattgctat ctaccggcat tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt 1381 atactcccac atatgccaga ttcagcaacg gatacggctt ccccaacttg cccacttcca 1441 tacgtgtcct ccttaccaga aatttatcct taaggtcgtc agctatcctg caggcgatct 1501 ctcgatttcg atcaagacat tcctttaatg gtcttttctg gacaccacta ggggtcagaa 1561 gtagttcatc aaactttctt ccctccctaa tctcattggt taccttgggc tatcgaaact 1621 taattaacca gtcaagtcag ctacttggcg agatcgactt gtctgggttt cgactacgct 1681 cagaattgcg tcagtcaagt tcgatctggt ccttgctatt gcacccgttc tccgattacg 1741 agtttcattt aaatcatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc 1801 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc 1861 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga 1921 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt 1981 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg 2041 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc 2101 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg 2161 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc 2221 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg 2281 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc 2341 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct 2401 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt 2461 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa 2521 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa 2581 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgca 2641 tttaaatttc cgaactctcc aaggccctcg tcggaaaatc ttcaaacctt tcgtccgatc 2701 catcttgcag gctacctctc gaacgaacta tcgcaagtct cttggccggc cttgcgcctt 2761 ggctattgct tggcagcgcc tatcgccagg tattactcca atcccgaata tccgagatcg 2821 ggatcacccg agagaagttc aacctacatc ctcaatcccg atctatccga gatccgagga 2881 atatcgaaat cggggcgcgc ctggcctccg cgccgggttt tggcgcctcc cgcgggcgcc 2941 cccctcgtca cggcgagcgc tgccacgtca gacgaagggc gcaggagcgt cctgatcctt 3001 ccgcccggac gctcaggaca gcggcccgct gctcataaga ctcggcctta gaaccccagt 3061 atcagcagaa ggacatttta ggacgggact tgggtgactc tagggcactg gttttctttc 3121 cagagagcgg aacaggcgag gaaaagtagt cccttctcgg cgattctgcg gagggatctc 3181 cgtggggcgg tgaacgccga tgattatata aggacgcgcc gggtgtggca cagctagttc 3241 cgtcgcagcc gggatttggg tcgcggttct tgtttgtgga tcgctgtgat cgtcacttgg 3301 tgagtagcgg gctgctgggc tggccggggc tttcgtggcc gccgggccgc tcggtgggac 3361 ggaagcgtgt ggagagaccg ccaagggctg tagtctgggt ccgcgagcaa ggttgccctg 3421 aactgggggt tggggggagc gcagcaaaat ggcggctgtt cccgagtctt gaatggaaga 3481 cgcttgtgag gcgggctgtg aggtcgttga aacaaggtgg ggggcatggt gggcggcaag 3541 aacccaaggt cttgagccct tcgctaatgc gggaaagctc ttattcgggt gagatgggct 3601 gggcaccatc tggggaccct gacgtgaagt ttgtcactga ctggagaact cggtttgtcg 3661 tctgttgcgg gggcggcagt tatggcggtg ccgttgggca gtgcacccgt acctttggga 3721 gcgcgcgccc tcgtcgtgtc gtgacgtcac ccgttctgtt ggcttataat gcagggtggg 3781 gccacctgcc ggtaggtgtg cggtaggctt ttctccgtcg caggacgcag ggttcgggcc 3841 tagggtaggc tctcctgaat cgacaggcgc cggacctctg gtgaggggag ggataagtga 3901 ggcgtcagtt tctttggtcg gttttatgta cctatcttct taagtagctg aagctccggt 3961 tttgaactat gcgctcgggg ttggcgagtg tgttttgtga agttttttag gcaccttttg 4021 aaatgtaatc atttgggtca atatgtaatt ttcagtgtta gacttgtaaa ttgtccgcta 4081 aattctggcc gtttttggct tttttgttag acaacatgac cgagtacaag cccacggtgc 4141 gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg 4201 actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc 4261 tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg 4321 acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg 4381 ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga 4441 tggagggcct cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg 4501 gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg 4561 aggccgccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cgcaacctcc 4621 ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc 4681 gcacctggtg catgacccgc aagcccggtg cctaggctag tatgtaagcc tagtcttaga 4741 taataaaatc gctatccatc gaagatggat gtgtgttggt tttttgtgtg tgtaacgcta 4801 ggcgcgcctg gtgtaccgag aacgatcctc tcagtgcgag tctcgacgat ccatatcgtt 4861 gcttggcagt cagccagtcg gaatccagct tgggacccag gaagtccaat cgtcagatat 4921 tgtactcaag cctggtcacg gcagcgtacc gatctgttta aacctagata ttgatagtct 4981 gatcggtcaa cgtataatcg agtcctagct tttgcaaaca tctatcaaga gacaggatca 5041 gcaggaggct ttcgcatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg 5101 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa 5161 gatcagttgg gtgcgcgagt gggttacatc gaactggatc tcaacagcgg taagatcctt 5221 gagagttttc gccccgaaga acgctttcca atgatgagca cttttaaagt tctgctatgt 5281 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat 5341 tctcagaatg acttggttga gtattcacca gtcacagaaa agcatcttac ggatggcatg 5401 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta 5461 cttctgacaa cgattggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat 5521 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag 5581 cgtgacacca cgatgcctgt agcaatggca acaaccttgc gtaaactatt aactggcgaa 5641 ctacttactc tagcttcccg gcaacagttg atagactgga tggaggcgga taaagttgca 5701 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc 5761 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt 5821 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc 5881 gctgagatag gtgcctcact gattaagcat tggtaaccga ttctaggtgc attggcgcag 5941 aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa 6001 cggatacggc ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc 6061 cttaagatcc cgaatcgttt aaactcgact ctggctctat cgaatctccg tcgtttcgag 6121 cttacgcgaa cagccgtggc gctcatttgc tcgtcgggca tcgaatctcg tcagctatcg 6181 tcagcttacc tttttggcag cgatcgcggc tcccgacatc ttggaccatt agctccacag 6241 gtatcttctt ccctctagtg gtcataacag cagcttcagc tacctctcaa ttcaaaaaac 6301 ccctcaagac ccgtttagag gccccaaggg gttatgctat caatcgttgc gttacacaca 6361 caaaaaacca acacacatcc atcttcgatg gatagcgatt ttattatcta actgctgatc 6421 gagtgtagcc agatctagta atcaattacg gggtcattag ttcatagccc atatatggag 6481 ttccg //