LOCUS Exported 5946 bp ds-DNA circular SYN 03-JUN-2021 DEFINITION Baculoviral transfer vector to express SARS-CoV-2 papain-like protease domain (nsp3) in insect cells. ACCESSION . VERSION . KEYWORDS pLIB_3xFlag-His-PLpro (nsp3) (SARS-CoV-2) SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5946) AUTHORS Lim CT, Tan KW, Wu M, Ulferts R, Armstrong LA, Ozono E, Drury LS, Milligan JC, Zeisner TU, Zeng J, Weissmann F, Canal B, Binerva-Todd G, Howell M, O'Reilly N, Beale R, Kulathu Y, Labib K, Diffley JFX TITLE Identifying SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of Nsp3 Papain-like Protease JOURNAL bioRxiv https://www.biorxiv.org/content/10.1101/2021.04.07.438804v1 REFERENCE 2 (bases 1 to 5946) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 3, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5946 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind 365..384 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 618..635 /label=L4440 /note="L4440 vector, forward primer" mobile_element 769..993 /mobile_element_type="transposon:Tn7" /note="mini-Tn7 element (right end of the Tn7 transposon)" CDS complement(1060..1593) /codon_start=1 /gene="aacC1" /product="gentamycin acetyltransferase" /label=GmR /note="confers resistance to gentamycin" /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR EEVMHFDIDPSTAT" promoter complement(1782..1810) /gene="intI1 (promoter lies within the coding sequence)" /label=Pc promoter /note="class 1 integron promoter" protein_bind 2206..2239 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." promoter 2287..2378 /gene="polh from Autographa californica" /label=polyhedrin promoter /note="promoter for the baculovirus polyhedrin gene" primer_bind 2303..2320 /label=Polyhedrin-F /note="Polyhedrin promoter, forward primer" regulatory 2422..2431 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2431..2496 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS 2503..2520 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 3613..3747 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3650..3669) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 3704..3723 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" mobile_element 3857..4022 /mobile_element_type="transposon:Tn7" /note="mini-Tn7 element (left end of the Tn7 transposon)" rep_origin 4206..4660 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4293..4312) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4502..4523 /label=F1ori-F /note="F1 origin, forward primer" promoter 4686..4790 /gene="bla" /label=AmpR promoter CDS 4791..5651 /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(5009..5028) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5822..464 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" ORIGIN 1 gagcgcagat accaaatact attcttctag tgtagccgta gttaggccac cacttcaaga 61 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca 121 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc 181 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca 241 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa 301 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc 361 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc 421 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg 481 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat 541 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca 601 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt 661 attttctcct tacgcatctg tgcggtattt cacaccgcat agaccagccg cgtaacctgg 721 caaaatcggt tacggttgag taataaatgg atgccctgcg taagcgggtg tgggcggaca 781 ataaagtctt aaactgaaca aaatagatct aaactatgac aataaagtct taaactagac 841 agaatagttg taaactgaaa tcagtccagt tatgctgtga aaaagcatac tggacttttg 901 ttatggctaa agcaaactct tcattttctg aagtgcaaat tgcccgtcgt attaaagagg 961 ggcgtggcca agggcatggt aaagactata ttcgcggcgt tgtgacaatt taccgaacaa 1021 ctccgcggcc gggaagccga tctcggcttg aacgaattgt taggtggcgg tacttgggtc 1081 gatatcaaag tgcatcactt cttcccgtat gcccaacttt gtatagagag ccactgcggg 1141 atcgtcaccg taatctgctt gcacgtagat cacataagca ccaagcgcgt tggcctcatg 1201 cttgaggaga ttgatgagcg cggtggcaat gccctgcctc cggtgctcgc cggagactgc 1261 gagatcatag atatagatct cactacgcgg ctgctcaaac ttgggcagaa cgtaagccgc 1321 gagagcgcca acaaccgctt cttggtcgaa ggcagcaagc gcgatgaatg tcttactacg 1381 gagcaagttc ccgaggtaat cggagtccgg ctgatgttgg gagtaggtgg ctacgtctcc 1441 gaactcacga ccgaaaagat caagagcagc ccgcatggat ttgacttggt cagggccgag 1501 cctacatgtg cgaatgatgc ccatacttga gccacctaac tttgttttag ggcgactgcc 1561 ctgctgcgta acatcgttgc tgctgcgtaa catcgttgct gctccataac atcaaacatc 1621 gacccacggc gtaacgcgct tgctgcttgg atgcccgagg catagactgt acaaaaaaac 1681 agtcataaca agccatgaaa accgccactg cgccgttacc accgctgcgt tcggtcaagg 1741 ttctggacca gttgcgtgag cgcatacgct acttgcatta cagtttacga accgaacagg 1801 cttatgtcaa ctgggtttgt gccttcatcc gtttccacgg tgtgcgtcac ccggcaacct 1861 tgggcagcag cgaagtcgag gcatttctgt cctggctggc gaacgagcgc aaggtttcgg 1921 tctccacgca tcgtcaggca ttggcggcct tgctgttctt ctacggcaag gtgctgtgca 1981 cggatctgcc cttgcttcag gagatcggta gacctcggcc gtcgcggcgc ttgccggtgg 2041 tgctgacccc ggatgaagtg gttcgcatcc tcggttttct ggaaggcgag catcgtttgt 2101 tcgcccagga ctctagctat agttctagtg gttggctaca gctttgtttg tactatcaac 2161 aggttgaact gctgatcaac agatcctcta cgcggccgcg gtaccataac ttcgtatagc 2221 atacattata cgaagttatc tggttttact agtatcgatt cgcgacctac tccggaatat 2281 taatagatca tggagataat taaaatgata accatctcgc aaataaataa gtattttact 2341 gttttcgtaa cagttttgta ataaaaaaac ctataaatat tccggattat tcataccgtc 2401 ccaccatcgg gcgcggatcc agccaccatg gactacaagg accacgacgg cgattacaag 2461 gatcatgaca tcgactacaa agacgacgat gacaagggtt cccaccatca ccaccatcac 2521 tctgctgtgc tgcaggaagt gcgtactatc aaggtgttca ccaccgtcga caacatcaac 2581 ctgcacaccc aggtggtcga catgtccatg acctacggcc agcagttcgg tcccacctac 2641 ttggacggtg ctgatgtgac caagatcaag ccccacaact cccacgaggg caagaccttc 2701 tacgtgctgc ccaacgacga taccctgcgt gtcgaggctt tcgagtacta ccatacaact 2761 gacccatcct tcctgggccg ttacatgtcc gctctgaacc acaccaagaa gtggaagtac 2821 cctcaagtga acggactgac ctccatcaag tgggctgaca acaactgcta cctcgctacc 2881 gctctgctga ccttgcagca gatcgagctg aagttcaacc ctccagctct gcaggacgct 2941 tactacagag ccagagctgg cgaggctgct aacttctgcg ctctgatcct ggcttactgc 3001 aacaagaccg tgggagagct gggagatgtg cgtgaaacca tgtcttacct gttccagcac 3061 gctaacctgg actcttgcaa gcgtgtcctg aacgtcgtgt gcaagacctg cggtcaacag 3121 cagactaccc tgaagggtgt cgaagctgtg atgtacatgg gcaccctgtc ctacgagcag 3181 ttcaagaagg gtgttcagat cccctgcact tgcggcaagc aggccactaa gtacctggtg 3241 cagcaagagt ctcccttcgt gatgatgtct gctcctcctg ctcaatacga actcaagcac 3301 ggaaccttca cttgcgcttc cgagtacacc ggcaactacc agtgcggtca ctacaagcac 3361 atcacctcca aagaaaccct gtactgcatc gacggtgccc tgctgaccaa gtcctctgag 3421 tacaaaggtc ccatcaccga cgtgttctac aaagagaact cctacacgac caccatcaag 3481 taaaagcttg tcgagaagta ctagaggatc ataatcagcc ataccacatt tgtagaggtt 3541 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca 3601 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc 3661 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc 3721 atcaatgtat cttatcatgt ctggatctga tcactgcttg agcctagaag atccggctgc 3781 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactatcata 3841 acccctagga gatccgaacc agataagtga aatctagttc caaactattt tgtcattttt 3901 aattttcgta ttagcttacg acgctacacc cagttcccat ctattttgtc actcttccct 3961 aaataatcct taaaaactcc atttccaccc ctcccagttc ccaactattt tgtccgccca 4021 cagcggggca tttttcttcc tgttatgttt ttaatcaaac atcctgccaa ctccatgtga 4081 caaaccgtca tcttcggcta ctttttctct gtcacagaat gaaaattttt ctgtcatctc 4141 ttcgttatta atgtttgtaa ttgactgaat atcaacgctt atttgcagcc tgaatggcga 4201 atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt 4261 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct 4321 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg 4381 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag 4441 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttcttaat 4501 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat 4561 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa 4621 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa 4681 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca 4741 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc 4801 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc 4861 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt 4921 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt 4981 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg 5041 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact 5101 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg 5161 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga 5221 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg 5281 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa 5341 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac 5401 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc 5461 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca 5521 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga 5581 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta 5641 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc 5701 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc 5761 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt 5821 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac 5881 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct 5941 tcagca //