LOCUS Exported 7437 bp ds-DNA circular SYN 20-OCT-2022 DEFINITION express SARS2-NSP1-mut4. ACCESSION . VERSION . KEYWORDS pVPSBA01-SARS-CoV2-nsp1 mutant4 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7437) AUTHORS Yuan S, Peng L, Park JJ, Hu Y, Devarkar SC, Dong MB, Shen Q, Wu S, Chen S, Lomakin IB, Xiong Y TITLE Nonstructural Protein 1 of SARS-CoV-2 Is a Potent Pathogenicity Factor Redirecting Host Protein Synthesis Machinery toward Viral RNA. JOURNAL Mol Cell. 2020 Dec 17;80(6):1055-1066.e6. doi: 10.1016/j.molcel.2020.10.034. Epub 2020 Oct 29. PUBMED 33188728 REFERENCE 2 (bases 1 to 7437) AUTHORS . TITLE Direct Submission JOURNAL Exported Oct 20, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7437 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(44..63) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" promoter 581..792 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 821..830 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1385..1408 /codon_start=1 /product="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" /label=Strep-Tag II /translation="WSHPQFEK" CDS 1445..1468 /codon_start=1 /product="peptide that binds Strep-Tactin(R), an engineered form of streptavidin" /label=Strep-Tag II /translation="WSHPQFEK" misc_feature 1480..2052 /label=IRES /note="internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)" primer_bind complement(1646..1663) /label=IRES reverse /note="IRES internal ribosome entry site, reverse primer. Also called pCDH-rev" primer_bind 1873..1892 /label=IRES-F /note="IRES internal ribosome entry site, forward primer" CDS 2085..2684 /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(2085..2104) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 2581..2601 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 2691..2739 /note="synthetic polyadenylation signal" primer_bind complement(3028..3045) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 3034..3258 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 3304..3732 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(3391..3410) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 3601..3622 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(3741..3761) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 3746..4075 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 3926..4061 /label=SV40 ori /note="SV40 origin of replication" primer_bind 3988..4007 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" CDS 4142..4936 /codon_start=1 /gene="aph(3')-II (or nptII)" /product="aminoglycoside phosphotransferase from Tn5" /label=NeoR/KanR /note="confers resistance to neomycin, kanamycin, and G418 (Geneticin(R))" /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF" primer_bind complement(4196..4215) /label=Neo-R /note="Neomycin resistance gene, reverse primer" primer_bind 4806..4825 /label=Neo-F /note="Neomycin resistance gene, forward primer" polyA_signal 5110..5231 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(5147..5166) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 5201..5220 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(5280..5296) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(5280..5296) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(5293..5315) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 5304..5320 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /note="The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-beta-D-thiogalactopyranoside (IPTG)." promoter complement(5328..5358) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 5373..5394 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(5511..5528) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(5682..6270) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(5762..5781) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(6441..7301) /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 7064..7083 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(7302..7406) /gene="bla" /label=AmpR promoter ORIGIN 1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg 61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg 121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc 181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttctagac 241 tatacagttg aagtcggaag tttacataca cttaagttgg agtcattaaa actcgttttt 301 caactactcc acaaatttct tgttaacaaa caatagtttt ggcaagtcag ttaggacatc 361 tactttgtgc atgacacaag tcatttttcc aacaattgtt tacagacaga ttatttcact 421 tataattcac tgtatcacaa ttccagtggg tcagaagttt acatacacta agttgactgt 481 gcctttaaac agcttggaaa attccagaaa atgatgtcat ggctttagag aggatcctag 541 gtcttgaaag gagtgggaat tggctccggt gcccgtcagt gggcagagcg cacatcgccc 601 acagtccccg agaagttggg gggaggggtc ggcaattgat ccggtgccta gagaaggtgg 661 cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg 721 ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc 781 gccagaacac aggaccggtg gtacccgtac ggccaccgcc gccaccatgg agagtcttgt 841 acccggcttt aacgagaaaa cacatgttca attgtcattg cctgtgctgc aagtccggga 901 cgtactggtt agggggttcg gagattcagt agaagaggtg ctgtctgagg cgaggcaaca 961 tttgaaagac gggacgtgcg ggctggttga ggtggaaaag ggagtacttc cacagttgga 1021 acaaccgtat gtttttatta agcggagtga tgccaggact gcgccgcacg gtcatgtcat 1081 ggttgagctt gtcgcagagc ttgaaggcat tcaatatggg cgctccggtg aaactcttgg 1141 agtacttgtg ccgcacgtag gcgaaattcc agtggcatat cggaaggtgc tgttgagaaa 1201 aaatgggtct gagggcgcag ggggccacag ttatggagcc gacctcaaat cctttgacct 1261 tggcgacgag ttggggacag atccctacga ggatttccag gaaaactgga atacgaaaca 1321 tagcagtggc gttactcgcg agttgatgcg cgaactgaac ggcggcctcg aaggcggcgg 1381 gggatggagc catccacaat tcgagaaagg cggtggttca ggaggaggta gcgggggtgg 1441 atcatggtca catccgcagt ttgaaaagta aggatcccgc ccctctccct ccccccccct 1501 aacgttactg gccgaagccg cttggaataa ggccggtgtg cgtttgtcta tatgttattt 1561 tccaccatat tgccgtcttt tggcaatgtg agggcccgga aacctggccc tgtcttcttg 1621 acgagcattc ctaggggtct ttcccctctc gccaaaggaa tgcaaggtct gttgaatgtc 1681 gtgaaggaag cagttcctct ggaagcttct tgaagacaaa caacgtctgt agcgaccctt 1741 tgcaggcagc ggaacccccc acctggcgac aggtgcctct gcggccaaaa gccacgtgta 1801 taagatacac ctgcaaaggc ggcacaaccc cagtgccacg ttgtgagttg gatagttgtg 1861 gaaagagtca aatggctctc ctcaagcgta ttcaacaagg ggctgaagga tgcccagaag 1921 gtaccccatt gtatgggatc tgatctgggg cctcggtgca catgctttac atgtgtttag 1981 tcgaggttaa aaaaacgtct aggccccccg aaccacgggg acgtggtttt cctttgaaaa 2041 acacgatgat aagcttgcca caacccacaa ggagacgacc ttccatgacc gagtacaagc 2101 ccacggtgcg cctcgccacc cgcgacgacg tcccccgggc cgtacgcacc ctcgccgccg 2161 cgttcgccga ctaccccgcc acgcgccaca ccgtcgaccc ggaccgccac atcgagcggg 2221 tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg 2281 tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg 2341 cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc 2401 agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg 2461 ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc 2521 ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc 2581 gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg 2641 aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctagctcgag aataaaagat 2701 ctttattttc attagatctg tgtgttggtt ttttgtgtga attttgagtg tatgtaaact 2761 tctgacccac tgggaatgtg atgaaagaaa taaaagctga aatgaatcat tctctctact 2821 attattctga tatttcacat tcttaaaata aagtggtgat cctaactgac ctaagacagg 2881 gaatttttac taggattaaa tgtcaggaat tgtgaaaaag tgagtttaaa tgtatttggc 2941 taaggtgtat gtaaacttcc gacttcaact gtataggcat gcggtaacca cgtgcggacc 3001 gaagcttaag tttaaaccgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg 3061 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt 3121 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg 3181 gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg catgctgggg 3241 atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct agggggtatc 3301 cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga 3361 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg 3421 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat 3481 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg 3541 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata 3601 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt 3661 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat 3721 ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc 3781 cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa 3841 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac 3901 catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc 3961 tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg cctctgcctc 4021 tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt gcaaaaagct 4081 cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag gatcgtttcg 4141 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt 4201 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc 4261 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact 4321 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt 4381 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca 4441 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat 4501 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg 4561 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga 4621 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga 4681 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa 4741 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga 4801 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt 4861 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct 4921 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac 4981 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc 5041 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc 5101 gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca 5161 aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc 5221 aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg 5281 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc 5341 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg 5401 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc 5461 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact 5521 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta 5581 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag 5641 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc 5701 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta 5761 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg 5821 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc 5881 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac 5941 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac 6001 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg 6061 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga 6121 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt 6181 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag 6241 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct 6301 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg 6361 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat 6421 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc 6481 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg 6541 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct 6601 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca 6661 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg 6721 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg 6781 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc 6841 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag 6901 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg 6961 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag 7021 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat 7081 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg 7141 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca 7201 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca 7261 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat 7321 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag 7381 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc //