LOCUS Exported 7622 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 endoRNAse. ACCESSION . VERSION . KEYWORDS pGBW-m4134206 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7622) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7622) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7622 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 1046..1066 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1067..1093 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" polyA_signal 1162..1283 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1173..1192) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1227..1246 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1398..1444 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1474..1610 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1655..1726 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1952..2540) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2032..2051) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 3040..4249 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 4086..4105 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 4253..5281 /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 5308..5356 /note="synthetic polyadenylation signal" CDS 5622..6482 /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(5840..5859) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 6495..6631 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6776..6847 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6852..6898 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6916..6964 /note="synthetic polyadenylation signal" enhancer 7045..7348 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 7350..7552 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 7503..7523 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgtca ctggagaacg ttgctttcaa tgtagtgaac aaggggcact ttgacggaca 61 gcaaggcgaa gtgccggtca gtatcataaa taatacggtg tacaccaaag tcgatggtgt 121 tgacgtggag ttatttgaaa acaagacaac tttgcctgtc aatgtggcct tcgagctgtg 181 ggcaaaacgc aacattaagc cagtacccga agtgaaaatc ctcaacaatc ttggcgttga 241 tattgccgca aacactgtta tctgggatta taagcgagat gcgcctgccc atatatccac 301 cattggagtg tgttctatga cagacatcgc taaaaagccc accgagacaa tttgcgcgcc 361 actcacggta ttctttgacg ggagggtcga cggtcaggtg gatctgttta gaaatgcccg 421 taacggcgta cttattactg aaggaagcgt gaaaggccta cagcccagtg ttgggccgaa 481 gcaagcaagc ctaaatggag tcacactgat cggagaggct gtgaaaaccc agttcaacta 541 ctataagaag gtcgacggcg tggttcagca gctgcctgag acatacttta cccaatcgcg 601 gaatctgcaa gaatttaagc ccagatccca gatggagatt gacttcttgg aactggccat 661 ggatgagttc atcgaacggt ataagctcga gggctacgcc ttcgagcaca tagtttatgg 721 ggatttttct cattcacagt taggcggtct gcacttgctc atcggattag cgaaacgctt 781 taaagagagc ccattcgaac ttgaagactt tatcccaatg gactcaactg tgaagaatta 841 cttcataact gacgctcaga cagggtccag caagtgcgtc tgttctgtta ttgatttgct 901 acttgatgat ttcgtggaga ttattaaatc acaggacctg tccgtggttt cgaaggtcgt 961 gaaagtaacc atcgactata cggaaattag tttcatgctt tggtgtaagg atggccatgt 1021 ggagacattt tatcctaagc tccaggaaaa cctttacttt cagtcgtacc cctatgacgt 1081 accagattat gcgtaaggat ccgcaggcct ctgctagctt gactgactga gatacagcgt 1141 accttcagct cacagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa 1201 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca 1261 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc 1321 agggggaggt gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtattggcc 1381 catctctatc ggtatcgtag cataacccct tggggcctct aaacgggtct tgaggggttt 1441 tttgtgcccc tcgggccgga ttgctatcta ccggcattgg cgcagaaaaa aatgcctgat 1501 gcgacgctgc gcgtcttata ctcccacata tgccagattc agcaacggat acggcttccc 1561 caacttgccc acttccatac gtgtcctcct taccagaaat ttatccttaa ggtcgtcagc 1621 tatcctgcag gcgatctctc gatttcgatc aagacattcc tttaatggtc ttttctggac 1681 accactaggg gtcagaagta gttcatcaaa ctttcttccc tccctaatct cattggttac 1741 cttgggctat cgaaacttaa ttaaccagtc aagtcagcta cttggcgaga tcgacttgtc 1801 tgggtttcga ctacgctcag aattgcgtca gtcaagttcg atctggtcct tgctattgca 1861 cccgttctcc gattacgagt ttcatttaaa tcatgtgagc aaaaggccag caaaaggcca 1921 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc 1981 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc 2041 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg 2101 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta 2161 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg 2221 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac 2281 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag 2341 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat 2401 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat 2461 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc 2521 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt 2581 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct 2641 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt 2701 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc 2761 gttcatccat agttgcattt aaatttccga actctccaag gccctcgtcg gaaaatcttc 2821 aaacctttcg tccgatccat cttgcaggct acctctcgaa cgaactatcg caagtctctt 2881 ggccggcctt gcgccttggc tattgcttgg cagcgcctat cgccaggtat tactccaatc 2941 ccgaatatcc gagatcggga tcacccgaga gaagttcaac ctacatcctc aatcccgatc 3001 tatccgagat ccgaggaata tcgaaatcgg ggcgcgcctg gcctccgcgc cgggttttgg 3061 cgcctcccgc gggcgccccc ctcgtcacgg cgagcgctgc cacgtcagac gaagggcgca 3121 ggagcgtcct gatccttccg cccggacgct caggacagcg gcccgctgct cataagactc 3181 ggccttagaa ccccagtatc agcagaagga cattttagga cgggacttgg gtgactctag 3241 ggcactggtt ttctttccag agagcggaac aggcgaggaa aagtagtccc ttctcggcga 3301 ttctgcggag ggatctccgt ggggcggtga acgccgatga ttatataagg acgcgccggg 3361 tgtggcacag ctagttccgt cgcagccggg atttgggtcg cggttcttgt ttgtggatcg 3421 ctgtgatcgt cacttggtga gtagcgggct gctgggctgg ccggggcttt cgtggccgcc 3481 gggccgctcg gtgggacgga agcgtgtgga gagaccgcca agggctgtag tctgggtccg 3541 cgagcaaggt tgccctgaac tgggggttgg ggggagcgca gcaaaatggc ggctgttccc 3601 gagtcttgaa tggaagacgc ttgtgaggcg ggctgtgagg tcgttgaaac aaggtggggg 3661 gcatggtggg cggcaagaac ccaaggtctt gagcccttcg ctaatgcggg aaagctctta 3721 ttcgggtgag atgggctggg caccatctgg ggaccctgac gtgaagtttg tcactgactg 3781 gagaactcgg tttgtcgtct gttgcggggg cggcagttat ggcggtgccg ttgggcagtg 3841 cacccgtacc tttgggagcg cgcgccctcg tcgtgtcgtg acgtcacccg ttctgttggc 3901 ttataatgca gggtggggcc acctgccggt aggtgtgcgg taggcttttc tccgtcgcag 3961 gacgcagggt tcgggcctag ggtaggctct cctgaatcga caggcgccgg acctctggtg 4021 aggggaggga taagtgaggc gtcagtttct ttggtcggtt ttatgtacct atcttcttaa 4081 gtagctgaag ctccggtttt gaactatgcg ctcggggttg gcgagtgtgt tttgtgaagt 4141 tttttaggca ccttttgaaa tgtaatcatt tgggtcaata tgtaattttc agtgttagac 4201 ttgtaaattg tccgctaaat tctggccgtt tttggctttt ttgttagaca acatgggtaa 4261 aaagcctgaa ctcaccgcga cgtctgtcga gaagtttctg atcgaaaagt tcgacagcgt 4321 ctccgacctg atgcagctct cggagggcga agaatctcgt gctttcagct tcgatgtagg 4381 agggcgtgga tatgtcctgc gggtaaatag ctgcgccgat ggtttctaca aagatcgtta 4441 tgtttatcgg cactttgcat ccgccgcgct cccgattccg gaagtgcttg acattgggga 4501 gttcagcgag agcctgacct attgcatctc ccgccgtgca cagggtgtca cgttgcaaga 4561 cctgcctgaa accgaactgc ccgctgttct gcagccggtc gcggaggcaa tggatgccat 4621 cgctgccgcc gatcttagcc agacgagcgg gttcggccca ttcggaccgc aaggaatcgg 4681 tcaatacact acatggcgtg atttcatatg cgcgattgct gatccccatg tgtatcactg 4741 gcaaactgtg atggacgaca ccgtcagtgc gtccgtcgcg caggctctcg atgagctgat 4801 gctttgggcc gaggactgcc ccgaagtccg gcacctcgtg cacgcggatt tcggctccaa 4861 caatgtcctg acggacaatg gccgcataac agcggtcatt gactggagcg aggcgatgtt 4921 cggggattcc caatacgagg tcgccaacat cttcttctgg aggccgtggt tggcttgtat 4981 ggagcagcag acgcgctact tcgagcggag gcatccggag cttgcaggat cgccccggct 5041 ccgggcgtat atgctccgca ttggtcttga ccaactctat cagagcttgg ttgacggcaa 5101 tttcgatgat gcagcttggg cgcagggtcg atgcgacgca atcgtccgat ccggagccgg 5161 gactgtcggg cgtacacaaa tcgcccgcag aagcgccgcc gtctggaccg atggctgtgt 5221 agaagtactc gccgatagtg gaaaccgacg ccccagcact cgtccgaggg caaaggaata 5281 agctagtatg taagcctagt cttagataat aaaatcgcta tccatcgaag atggatgtgt 5341 gttggttttt tgtgtgtgta acgctaggcg cgcctggtgt accgagaacg atcctctcag 5401 tgcgagtctc gacgatccat atcgttgctt ggcagtcagc cagtcggaat ccagcttggg 5461 acccaggaag tccaatcgtc agatattgta ctcaagcctg gtcacggcag cgtaccgatc 5521 tgtttaaacc tagatattga tagtctgatc ggtcaacgta taatcgagtc ctagcttttg 5581 caaacatcta tcaagagaca ggatcagcag gaggctttcg catgagtatt caacatttcc 5641 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa 5701 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc gcgagtgggt tacatcgaac 5761 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgc tttccaatga 5821 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag 5881 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtat tcaccagtca 5941 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca 6001 tgagtgataa cactgcggcc aacttacttc tgacaacgat tggaggaccg aaggagctaa 6061 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc 6121 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa 6181 ccttgcgtaa actattaact ggcgaactac ttactctagc ttcccggcaa cagttgatag 6241 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct 6301 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac 6361 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa 6421 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt 6481 aaccgattct aggtgcattg gcgcagaaaa aaatgcctga tgcgacgctg cgcgtcttat 6541 actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc cacttccata 6601 cgtgtcctcc ttaccagaaa tttatcctta agatcgttta aactcgactc tggctctatc 6661 gaatctccgt cgtttcgagc ttacgcgaac agccgtggcg ctcatttgct cgtcgggcat 6721 cgaatctcgt cagctatcgt cagcttacct ttttggcagc gatcgcggct cccgacatct 6781 tggaccatta gctccacagg tatcttcttc cctctagtgg tcataacagc agcttcagct 6841 acctctcaat tcaaaaaacc cctcaagacc cgtttagagg ccccaagggg ttatgctatc 6901 aatcgttgcg ttacacacac aaaaaaccaa cacacatcca tcttcgatgg atagcgattt 6961 tattatctaa ctgctgatcg agtgtagcca gatctagtaa tcaattacgg ggtcattagt 7021 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg 7081 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc 7141 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc 7201 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg 7261 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat 7321 ctacgtatta gtcatcgcta ttaccatgct gatgcggttt tggcagtaca tcaatgggcg 7381 tggatagcgg tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag 7441 tttgttttgg caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt 7501 gacgcaaatg ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt 7561 gaaccgtcag atcagatctt tgtcgatcct accatccact cgacacaccc gccagcggcc 7621 gc //