LOCUS Exported 5184 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp10. ACCESSION . VERSION . KEYWORDS pGBW-m4134133 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5184) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5184) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5184 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 425..445 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 479..556 /codon_start=1 /product="calmodulin-binding peptide" /label=CBP /note="derived from skeletal muscle myosin light chain kinase; binds calmodulin with nanomolar affinity in the presence of calcium" /translation="KRRWKKNFIAVSAANRFKKISSSGAL" CDS 584..604 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" polyA_signal 1053..1174 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1064..1083) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1118..1137 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1289..1335 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1365..1501 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1546..1617 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1843..2431) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1923..1942) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 3177..4037 /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(3395..3414) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 4050..4186 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 4338..4409 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4414..4460 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 4478..4526 /note="synthetic polyadenylation signal" enhancer 4607..4910 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4912..5114 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 5065..5085 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 5183..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggca gggaatgcga cagaggttcc tgccaactca acggtcttgt ctttttgtgc 61 tttcgccgtg gacgctgcaa aagcctacaa ggattatcta gcctcgggtg gacagccaat 121 cactaactgc gtaaagatgc tttgtaccca caccggcact ggccaagcaa ttacagtgac 181 ccccgaagct aatatggacc aggagtcctt cggaggtgcg agctgctgcc tgtactgtcg 241 ttgtcatata gatcacccga atcccaaagg gttttgcgac ctcaagggca aatatgtcca 301 gatcccaacg acatgcgcca acgatcctgt gggattcact ctgaagaaca ccgtatgtac 361 cgtttgcggg atgtggaagg ggtatggctg tagttgcgac caattacggg aaccgatgct 421 gcaagaaaac ctttactttc agtcgggacg gcgaatcccg gggctgataa atccctggaa 481 aaggagatgg aagaagaact ttattgcagt gtctgcggct aaccggttca aaaagatcag 541 ttcaagcggc gccctagatt acgacattcc tacgaccgcc tccgagaatc tctatttcca 601 gggtgaattt ggcttggcac aacatgacga ggccgtcgat aacaagttta ataaagaaca 661 gcagaacgct ttctatgaga tccttcactt accaaatctg aacgaagagc aacgcaacgc 721 tttcattcag agcctgaaag acgatccctc gcagtctgca aatctcctgg ccgaagccaa 781 gaagctgaat gacgcacagg ctccaaaagt tgataacaag tttaacaagg agcagcagaa 841 tgctttctac gagatattgc accttcctaa tctgaatgag gaacaaagaa atgcatttat 901 ccagtcccta aaagacgatc catcacagag tgccaacctg cttgcggaag ccaaaaagct 961 caatgatgcc caggccccga aagtagacgc gaaccatcaa taagctagct tgactgactg 1021 agatacagcg taccttcagc tcacagacat gataagatac attgatgagt ttggacaaac 1081 cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt 1141 atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca ttcattttat 1201 gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg 1261 tggtattggc ccatctctat cggtatcgta gcataacccc ttggggcctc taaacgggtc 1321 ttgaggggtt ttttgtgccc ctcgggccgg attgctatct accggcattg gcgcagaaaa 1381 aaatgcctga tgcgacgctg cgcgtcttat actcccacat atgccagatt cagcaacgga 1441 tacggcttcc ccaacttgcc cacttccata cgtgtcctcc ttaccagaaa tttatcctta 1501 aggtcgtcag ctatcctgca ggcgatctct cgatttcgat caagacattc ctttaatggt 1561 cttttctgga caccactagg ggtcagaagt agttcatcaa actttcttcc ctccctaatc 1621 tcattggtta ccttgggcta tcgaaactta attaaccagt caagtcagct acttggcgag 1681 atcgacttgt ctgggtttcg actacgctca gaattgcgtc agtcaagttc gatctggtcc 1741 ttgctattgc acccgttctc cgattacgag tttcatttaa atcatgtgag caaaaggcca 1801 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc 1861 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact 1921 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct 1981 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag 2041 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca 2101 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa 2161 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc 2221 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag 2281 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg 2341 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca 2401 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc 2461 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag 2521 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata 2581 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat 2641 ctgtctattt cgttcatcca tagttgcatt taaatttccg aactctccaa ggccctcgtc 2701 ggaaaatctt caaacctttc gtccgatcca tcttgcaggc tacctctcga acgaactatc 2761 gcaagtctct tggccggcct tgcgccttgg ctattgcttg gcagcgccta tcgccaggta 2821 ttactccaat cccgaatatc cgagatcggg atcacccgag agaagttcaa cctacatcct 2881 caatcccgat ctatccgaga tccgaggaat atcgaaatcg gggcgcgcct ggtgtaccga 2941 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc 3001 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac 3061 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc 3121 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga 3181 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt 3241 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag 3301 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag 3361 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta 3421 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg 3481 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca 3541 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag 3601 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc 3661 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg 3721 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc 3781 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg 3841 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg 3901 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga 3961 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac 4021 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga 4081 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac 4141 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc ccgaatcgtt 4201 taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg 4261 cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac ctttttggca 4321 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt 4381 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga 4441 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc 4501 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctagt 4561 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta 4621 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga 4681 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt 4741 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta 4801 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg 4861 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg ctgatgcggt 4921 tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc 4981 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat 5041 gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct 5101 atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc ctaccatcca 5161 ctcgacacac ccgccagcgg ccgc //