LOCUS Exported 7008 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp10. ACCESSION . VERSION . KEYWORDS pGBW-m4134407 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7008) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7008) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7008 /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" regulatory 577..586 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1003..1023 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1024..1050 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" polyA_signal 1119..1240 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1130..1149) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1184..1203 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1355..1401 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1431..1567 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1612..1683 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1909..2497) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1989..2008) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 2997..4206 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 4043..4062 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 4210..5238 /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 5265..5313 /note="synthetic polyadenylation signal" CDS 5579..6439 /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(5797..5816) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 6452..6588 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6740..6811 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6816..6862 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6880..6928 /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 ccatggcagg gaatgcgaca 601 gaggttcctg ccaactcaac ggtcttgtct ttttgtgctt tcgccgtgga cgctgcaaaa 661 gcctacaagg attatctagc ctcgggtgga cagccaatca ctaactgcgt aaagatgctt 721 tgtacccaca ccggcactgg ccaagcaatt acagtgaccc ccgaagctaa tatggaccag 781 gagtccttcg gaggtgcgag ctgctgcctg tactgtcgtt gtcatataga tcacccgaat 841 cccaaagggt tttgcgacct caagggcaaa tatgtccaga tcccaacgac atgcgccaac 901 gatcctgtgg gattcactct gaagaacacc gtatgtaccg tttgcgggat gtggaagggg 961 tatggctgta gttgcgacca attacgggaa ccgatgctgc aagaaaacct ttactttcag 1021 tcgtacccct atgacgtacc agattatgcg taaggatccg caggcctctg ctagcttgac 1081 tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg atgagtttgg 1141 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat 1201 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca 1261 ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt aaaacctcta 1321 caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg ggcctctaaa 1381 cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg gcattggcgc 1441 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc 1501 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta 1561 tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag acattccttt 1621 aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt tcttccctcc 1681 ctaatctcat tggttacctt gggctatcga aacttaatta accagtcaag tcagctactt 1741 ggcgagatcg acttgtctgg gtttcgacta cgctcagaat tgcgtcagtc aagttcgatc 1801 tggtccttgc tattgcaccc gttctccgat tacgagtttc atttaaatca tgtgagcaaa 1861 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct 1921 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac 1981 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc 2041 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc 2101 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg 2161 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga 2221 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag 2281 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta 2341 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag 2401 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg 2461 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac 2521 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc 2581 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag 2641 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc 2701 agcgatctgt ctatttcgtt catccatagt tgcatttaaa tttccgaact ctccaaggcc 2761 ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt gcaggctacc tctcgaacga 2821 actatcgcaa gtctcttggc cggccttgcg ccttggctat tgcttggcag cgcctatcgc 2881 caggtattac tccaatcccg aatatccgag atcgggatca cccgagagaa gttcaaccta 2941 catcctcaat cccgatctat ccgagatccg aggaatatcg aaatcggggc gcgcctggcc 3001 tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc gtcacggcga gcgctgccac 3061 gtcagacgaa gggcgcagga gcgtcctgat ccttccgccc ggacgctcag gacagcggcc 3121 cgctgctcat aagactcggc cttagaaccc cagtatcagc agaaggacat tttaggacgg 3181 gacttgggtg actctagggc actggttttc tttccagaga gcggaacagg cgaggaaaag 3241 tagtcccttc tcggcgattc tgcggaggga tctccgtggg gcggtgaacg ccgatgatta 3301 tataaggacg cgccgggtgt ggcacagcta gttccgtcgc agccgggatt tgggtcgcgg 3361 ttcttgtttg tggatcgctg tgatcgtcac ttggtgagta gcgggctgct gggctggccg 3421 gggctttcgt ggccgccggg ccgctcggtg ggacggaagc gtgtggagag accgccaagg 3481 gctgtagtct gggtccgcga gcaaggttgc cctgaactgg gggttggggg gagcgcagca 3541 aaatggcggc tgttcccgag tcttgaatgg aagacgcttg tgaggcgggc tgtgaggtcg 3601 ttgaaacaag gtggggggca tggtgggcgg caagaaccca aggtcttgag cccttcgcta 3661 atgcgggaaa gctcttattc gggtgagatg ggctgggcac catctgggga ccctgacgtg 3721 aagtttgtca ctgactggag aactcggttt gtcgtctgtt gcgggggcgg cagttatggc 3781 ggtgccgttg ggcagtgcac ccgtaccttt gggagcgcgc gccctcgtcg tgtcgtgacg 3841 tcacccgttc tgttggctta taatgcaggg tggggccacc tgccggtagg tgtgcggtag 3901 gcttttctcc gtcgcaggac gcagggttcg ggcctagggt aggctctcct gaatcgacag 3961 gcgccggacc tctggtgagg ggagggataa gtgaggcgtc agtttctttg gtcggtttta 4021 tgtacctatc ttcttaagta gctgaagctc cggttttgaa ctatgcgctc ggggttggcg 4081 agtgtgtttt gtgaagtttt ttaggcacct tttgaaatgt aatcatttgg gtcaatatgt 4141 aattttcagt gttagacttg taaattgtcc gctaaattct ggccgttttt ggcttttttg 4201 ttagacaaca tgggtaaaaa gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc 4261 gaaaagttcg acagcgtctc cgacctgatg cagctctcgg agggcgaaga atctcgtgct 4321 ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg taaatagctg cgccgatggt 4381 ttctacaaag atcgttatgt ttatcggcac tttgcatccg ccgcgctccc gattccggaa 4441 gtgcttgaca ttggggagtt cagcgagagc ctgacctatt gcatctcccg ccgtgcacag 4501 ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg 4561 gaggcaatgg atgccatcgc tgccgccgat cttagccaga cgagcgggtt cggcccattc 4621 ggaccgcaag gaatcggtca atacactaca tggcgtgatt tcatatgcgc gattgctgat 4681 ccccatgtgt atcactggca aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag 4741 gctctcgatg agctgatgct ttgggccgag gactgccccg aagtccggca cctcgtgcac 4801 gcggatttcg gctccaacaa tgtcctgacg gacaatggcc gcataacagc ggtcattgac 4861 tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg ccaacatctt cttctggagg 4921 ccgtggttgg cttgtatgga gcagcagacg cgctacttcg agcggaggca tccggagctt 4981 gcaggatcgc cccggctccg ggcgtatatg ctccgcattg gtcttgacca actctatcag 5041 agcttggttg acggcaattt cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc 5101 gtccgatccg gagccgggac tgtcgggcgt acacaaatcg cccgcagaag cgccgccgtc 5161 tggaccgatg gctgtgtaga agtactcgcc gatagtggaa accgacgccc cagcactcgt 5221 ccgagggcaa aggaataagc tagtatgtaa gcctagtctt agataataaa atcgctatcc 5281 atcgaagatg gatgtgtgtt ggttttttgt gtgtgtaacg ctaggcgcgc ctggtgtacc 5341 gagaacgatc ctctcagtgc gagtctcgac gatccatatc gttgcttggc agtcagccag 5401 tcggaatcca gcttgggacc caggaagtcc aatcgtcaga tattgtactc aagcctggtc 5461 acggcagcgt accgatctgt ttaaacctag atattgatag tctgatcggt caacgtataa 5521 tcgagtccta gcttttgcaa acatctatca agagacagga tcagcaggag gctttcgcat 5581 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt 5641 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcgcg 5701 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga 5761 agaacgcttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg 5821 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt 5881 tgagtattca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg 5941 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgattgg 6001 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga 6061 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc 6121 tgtagcaatg gcaacaacct tgcgtaaact attaactggc gaactactta ctctagcttc 6181 ccggcaacag ttgatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc 6241 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg 6301 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac 6361 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc 6421 actgattaag cattggtaac cgattctagg tgcattggcg cagaaaaaaa tgcctgatgc 6481 gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac ggcttcccca 6541 acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaaga tcccgaatcg 6601 tttaaactcg actctggctc tatcgaatct ccgtcgtttc gagcttacgc gaacagccgt 6661 ggcgctcatt tgctcgtcgg gcatcgaatc tcgtcagcta tcgtcagctt acctttttgg 6721 cagcgatcgc ggctcccgac atcttggacc attagctcca caggtatctt cttccctcta 6781 gtggtcataa cagcagcttc agctacctct caattcaaaa aacccctcaa gacccgttta 6841 gaggccccaa ggggttatgc tatcaatcgt tgcgttacac acacaaaaaa ccaacacaca 6901 tccatcttcg atggatagcg attttattat ctaactgctg atcgagtgta gccagatcta 6961 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccg //