LOCUS Exported 7319 bp ds-DNA circular SYN 03-JUN-2021 DEFINITION Baculoviral transfer vector to express nsp7-His6-nsp8 fusion protein (SARS-CoV-2) in insect cells. ACCESSION . VERSION . KEYWORDS pBIG1b_nsp7-His6-nsp8 (SARS-CoV-2) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7319) AUTHORS Bertolin AP, Weissmann F, Zeng J, Posse V, Milligan JC, Canal B, Ulferts R, Wu M, Drury LS, Howell M, Beale R, Diffley JFX TITLE Identifying SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of Nsp12/7/8 RNA-dependent RNA Polymerase JOURNAL bioRxiv https://www.biorxiv.org/content/10.1101/2021.04.07.438807v1 REFERENCE 2 (bases 1 to 7319) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 3, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7319 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 215..748 /codon_start=1 /gene="aacC1" /product="gentamycin acetyltransferase" /label=GmR /note="confers resistance to gentamycin" /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR EEVMHFDIDPSTAT" mobile_element 815..1039 /mobile_element_type="transposon:Tn7" /note="mini-Tn7 element (right end of the Tn7 transposon)" primer_bind complement(1173..1190) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(1344..1932) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1424..1443) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(2103..2963) /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 2726..2745 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(2964..3068) /gene="bla" /label=AmpR promoter rep_origin complement(3094..3548) /direction=LEFT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3231..3252) /label=F1ori-F /note="F1 origin, forward primer" primer_bind 3442..3461 /label=F1ori-R /note="F1 origin, reverse primer" CDS complement(3589..4380) /codon_start=1 /gene="aadA" /product="aminoglycoside adenylyltransferase (Murphy, 1985)" /label=SmR /note="confers resistance to spectinomycin and streptomycin" /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK" mobile_element 4904..5069 /mobile_element_type="transposon:Tn7" /note="mini-Tn7 element (left end of the Tn7 transposon)" polyA_signal 5384..5518 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5408..5427) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 5462..5481 /label=SV40pA-R /note="SV40 polyA, reverse primer" CDS complement(6245..6262) /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" promoter complement(6564..6655) /gene="polh from Autographa californica" /label=polyhedrin promoter /note="promoter for the baculovirus polyhedrin gene" primer_bind complement(6622..6639) /label=Polyhedrin-F /note="Polyhedrin promoter, forward primer" polyA_signal 6750..6798 /label=HSV TK poly(A) signal /note="herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)" primer_bind 6823..6842 /label=TK-pA-R /note="Thymidine kinase polyA, reverse primer" protein_bind complement(6888..6921) /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." promoter 7317..26 /gene="intI1 (promoter lies within the coding sequence)" /label=Pc promoter /note="class 1 integron promoter" ORIGIN 1 acataagcct gttcggttcg taaactgtaa tgcaagtagc gtatgcgctc acgcaactgg 61 tccagaacct tgaccgaacg cagcggtggt aacggcgcag tggcggtttt catggcttgt 121 tatgactgtt tttttgtaca gtctatgcct cgggcatcca agcagcaagc gcgttacgcc 181 gtgggtcgat gtttgatgtt atggagcagc aacgatgtta cgcagcagca acgatgttac 241 gcagcagggc agtcgcccta aaacaaagtt aggtggctca agtatgggca tcattcgcac 301 atgtaggctc ggccctgacc aagtcaaatc catgcgggct gctcttgatc ttttcggtcg 361 tgagttcgga gacgtagcca cctactccca acatcagccg gactccgatt acctcgggaa 421 cttgctccgt agtaagacat tcatcgcgct tgctgccttc gaccaagaag cggttgttgg 481 cgctctcgcg gcttacgttc tgcccaagtt tgagcagccg cgtagtgaga tctatatcta 541 tgatctcgca gtctccggcg agcaccggag gcagggcatt gccaccgcgc tcatcaatct 601 cctcaagcat gaggccaacg cgcttggtgc ttatgtgatc tacgtgcaag cagattacgg 661 tgacgatccc gcagtggctc tctatacaaa gttgggcata cgggaagaag tgatgcactt 721 tgatatcgac ccaagtaccg ccacctaaca attcgttcaa gccgagatcg gcttcccggc 781 cgcggagttg ttcggtaaat tgtcacaacg ccgcgaatat agtctttacc atgcccttgg 841 ccacgcccct ctttaatacg acgggcaatt tgcacttcag aaaatgaaga gtttgcttta 901 gccataacaa aagtccagta tgctttttca cagcataact ggactgattt cagtttacaa 961 ctattctgtc tagtttaaga ctttattgtc atagtttaga tctattttgt tcagtttaag 1021 actttattgt ccgcccacac ccgcttacgc agggcatcca tttattactc aaccgtaacc 1081 gattttgcca ggttacgcgg ctggtctatg cggtgtgaaa taccgcacag atgcgtaagg 1141 agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc 1201 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa 1261 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt 1321 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa 1381 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt 1441 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg 1501 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc 1561 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc 1621 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta 1681 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct 1741 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaatagt atttggtatc 1801 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa 1861 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa 1921 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa 1981 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt 2041 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac 2101 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc 2161 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc 2221 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata 2281 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc 2341 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc 2401 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca 2461 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa 2521 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca 2581 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt 2641 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt 2701 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg 2761 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga 2821 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc 2881 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg 2941 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag 3001 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg 3061 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa 3121 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca 3181 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga 3241 acaagagtcc actattaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca 3301 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg 3361 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc 3421 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc 3481 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca 3541 gggcgcgtcc cattcgccaa tccgtcggct tgaacgaatt gttagacatt atttgccgac 3601 taccttggtg atctcgcctt tcacgtagtg gacaaattct tccaactgat ctgcgcgcga 3661 ggccaagcga tcttcttctt gtccaagata agcctgtcta gcttcaagta tgacgggctg 3721 atactgggcc ggcaggcgct ccattgccca gtcggcagcg acatccttcg gcgcgatttt 3781 gccggttact gcgctgtacc aaatgcggga caacgtaagc actacatttc gctcatcgcc 3841 agcccagtcg ggcggcgagt tccatagcgt taaggtttca tttagcgcct caaatagatc 3901 ctgttcagga accggatcaa agagttcctc cgccgctgga cctaccaagg caacgctatg 3961 ttctcttgct tttgtcagca agatagccag atcaatgtcg atcgtggctg gctcgaagat 4021 acctgcaaga atgtcattgc gctgccattc tccaaattgc agttcgcgct tagctggata 4081 acgccacgga atgatgtcgt cgtgcacaac aatggtgact tctacagcgc ggagaatctc 4141 gctctctcca ggggaagccg aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt 4201 ttcatcaagc cttacggtca ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc 4261 gccatccact gcggagccgt acaaatgtac ggccagcaac gtcggttcga gatggcgctc 4321 gatgacgcca actacctctg atagttgagt cgatacttcg gcgatcaccg cttccctcat 4381 gatgtttaac tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctccataa 4441 catcaaacat cgacccacgg cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg 4501 taccccaaaa aaacagtcat aacaagccat gaaaaccgcc actgcgccgt taccaccgct 4561 gcgttcggtc aaggttctgg accagttgcg tgagcgcata cgctacttac cggtttattg 4621 actaccggaa gcagtgtgac cgtgtgcttc tcaaatgcct gaggccagtt tgctcaggct 4681 ctccccgtgg aggtaataat tgacgatatg atcatttatt ctgcctccca gttcaggctg 4741 caaataagcg ttgatattca gtcaattaca aacattaata acgaagagat gacagaaaaa 4801 ttttcattct gtgacagaga aaaagtagcc gaagatgacg gtttgtcaca tggagttggc 4861 aggatgtttg attaaaaaca taacaggaag aaaaatgccc cgctgtgggc ggacaaaata 4921 gttgggaact gggaggggtg gaaatggagt ttttaaggat tatttaggga agagtgacaa 4981 aatagatggg aactgggtgt agcgtcgtaa gctaatacga aaattaaaaa tgacaaaata 5041 gtttggaact agatttcact tatctggttc ggatctcctg tttaaacgtg caatagtatc 5101 cagtttttaa ttaacccgta gtggctatgg cagggcttgc cgccccgacg ttggctgcga 5161 gccctgggcc ttcacccgaa cttgggggtt ggggtgggga aaaggaagaa acgcgggcgt 5221 attggtccca atggggtctc ggtggggtat cgacgtaacc ccgattgaga tatagattta 5281 tttaaatggt tatgatagtt attgctcagc ggtggcagca gccaactcag cttcctttcg 5341 ggctttgtta gcagccggat cttctaggct caagcagtga tcagatccag acatgataag 5401 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg 5461 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa 5521 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta 5581 aagcaagtaa aacctctaca aatgtggtat ggctgattat gatcctctag tacttctcga 5641 caagctttta ctgcagcttc acagcggagt tagcacgcag agcagtcacg atcaggggcc 5701 aagccaggtt gggagagttg tccatggaga tctcggacag ctgcacgatc ttggagtcag 5761 cgtccaccac ttgctggatc tcccacagag cggaagcgta ggtgaaggtg gtgccgtcgc 5821 aagtgttctt gtaggtgttg tagtcgggga tgaccaccat cagcttagca gcggtggtca 5881 gagggatgat gttcaggggc acgcaaccgt cacgagcgtt gttgatgatg ttgttcagag 5941 cgtcgttgtc cagcttgcgg agcatggtga acagcatggt ctgcatagcg gaggtcactt 6001 tagcacgctt gtcctcggaa cgagcctgct tgtacatctg ggtcatagcc tggtcagcca 6061 tcttctcgag cttgcgctgc atagcagcgt cacggtcgaa ctcggacttg gccacgttca 6121 gggacttttt cagcttcttc agcaccacct cagagtcgcc gttagccaca gcctgctcgt 6181 aagcctcttg agcggtagcg aaagcagcgt aggagggcag agaggagaac tcggaagcga 6241 tagcgtgatg gtggtgatgg tgctgcaggg tagcacggtt gtccagcatc tcttcgcaca 6301 gcttgttgat gtccacagca ccttgcatag acagcaggac ggacagcagg gacaccatct 6361 tctcgaaagc ctcggtggtg tccttagcca gcaggatgtc gttgtgcagt tggacgcact 6421 gagcccacag cttgctggag gattccacac gcagctgttg cagcacagac agcagcacca 6481 cggaggtgca cttcacgtcg gacatcttgg acatggtggc tggatccgcg cccgatggtg 6541 ggacggtatg aataatccgg aatatttata ggttttttta ttacaaaact gttacgaaaa 6601 cagtaaaata cttatttatt tgcgagatgg ttatcatttt aattatctcc atgatctatt 6661 aatattccgg agtaggtcga tttaaatctt tagaccatag agcgttcact tgacagagtg 6721 ccagccctgg gaccgaaccc cgcgtttatg aacaaacgac ccaacacccg tgcgttttat 6781 tctgtctttt tattgccgtc atagcgcggg ttccttccgg tattgtctcc ttccgtgttt 6841 cagttagcct cgataaaaac cccgattgag atatagttta aacccagata acttcgtata 6901 atgtatgcta tacgaagtta tggtaccgcg gccgcgtaga ggatctgttg atcagcagtt 6961 caacctgttg atagtacaaa caaagctgta gccaaccact agaactatag ctagagtcct 7021 gggcgaacaa acgatgctcg ccttccagaa aaccgaggat gcgaaccact tcatccgggg 7081 tcagcaccac cggcaagcgc cgcgacggcc gaggtctacc gatctcctga agcaagggca 7141 gatccgtgca cagcaccttg ccgtagaaga acagcaaggc cgccaatgcc tgacgatgcg 7201 tggagaccga aaccttgcgc tcgttcgcca gccaggacag aaatgcctcg acttcgctgc 7261 tgcccaaggt tgccgggtga cgcacaccgt ggaaacggat gaaggcacaa acccagttg //