LOCUS Exported 7010 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp6. ACCESSION . VERSION . KEYWORDS pGBW-m4134157 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7010) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7010) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7010 /organism="synthetic DNA construct" /mol_type="other DNA" promoter 2..204 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 155..175 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 273..282 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1146..1166 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1167..1193 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" polyA_signal 1246..1367 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1257..1276) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1311..1330 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1482..1528 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1558..1694 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1739..1810 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2036..2624) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2116..2135) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 3124..4333 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 4170..4189 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 4337..4936 /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(4337..4356) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4833..4853 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 4963..5011 /note="synthetic polyadenylation signal" CDS 5277..6137 /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(5495..5514) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 6150..6286 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6438..6509 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6514..6560 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6578..6626 /note="synthetic polyadenylation signal" enhancer 6707..7010 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" ORIGIN 1 ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt 61 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac 121 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg 181 tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc tttgtcgatc 241 ctaccatcca ctcgacacac ccgccagcgg ccgccaccat ggcagtgaaa agaacaatca 301 agggtacaca ccactggttg ttactcacaa ttttgacttc acttttagtt ttagtccaga 361 gtactcaatg gtctttgttc ttttttttgt atgaaaatgc ctttttacct tttgctatgg 421 gtattattgc tatgtctgct tttgcaatga tgtttgtcaa acataagcat gcatttctct 481 gtttgttttt gttaccttct cttgccactg tagcttattt taatatggtc tatatgcctg 541 ctagttgggt gatgcgtatt atgacatggt tggatatggt tgatactagt ttgtctggtt 601 ttaagctaaa agactgtgtt atgtatgcat cagctgtagt gttactaatc cttatgacag 661 caagaactgt gtatgatgat ggtgctagga gagtgtggac acttatgaat gtcttgacac 721 tcgtttataa agtttattat ggtaatgctt tagatcaagc catttccatg tgggctctta 781 taatctctgt tacttctaac tactcaggtg tagttacaac tgtcatgttt ttggccagag 841 gtattgtttt tatgtgtgtt gagtattgcc ctattttctt cataactggt aatacacttc 901 agtgtataat gctagtttat tgtttcttag gctatttttg tacttgttac tttggcctct 961 tttgtttact caaccgctac tttagactga ctcttggtgt ttatgattac ttagtttcta 1021 cacaggagtt tagatatatg aattcacagg gactactccc acccaagaat agcatagatg 1081 ccttcaaact caacattaaa ttgttgggtg ttggtggcaa accttgtatc aaagtagcca 1141 ctgtagaaaa cctttacttt cagtcgtacc cctatgacgt accagattat gcgtaagcta 1201 gcttgactga ctgagataca gcgtaccttc agctcacaga catgataaga tacattgatg 1261 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg 1321 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt 1381 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa 1441 acctctacaa atgtggtatt ggcccatctc tatcggtatc gtagcataac cccttggggc 1501 ctctaaacgg gtcttgaggg gttttttgtg cccctcgggc cggattgcta tctaccggca 1561 ttggcgcaga aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca catatgccag 1621 attcagcaac ggatacggct tccccaactt gcccacttcc atacgtgtcc tccttaccag 1681 aaatttatcc ttaaggtcgt cagctatcct gcaggcgatc tctcgatttc gatcaagaca 1741 ttcctttaat ggtcttttct ggacaccact aggggtcaga agtagttcat caaactttct 1801 tccctcccta atctcattgg ttaccttggg ctatcgaaac ttaattaacc agtcaagtca 1861 gctacttggc gagatcgact tgtctgggtt tcgactacgc tcagaattgc gtcagtcaag 1921 ttcgatctgg tccttgctat tgcacccgtt ctccgattac gagtttcatt taaatcatgt 1981 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc 2041 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa 2101 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc 2161 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg 2221 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc 2281 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc 2341 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca 2401 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact 2461 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg 2521 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt 2581 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct 2641 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga 2701 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa 2761 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac 2821 ctatctcagc gatctgtcta tttcgttcat ccatagttgc atttaaattt ccgaactctc 2881 caaggccctc gtcggaaaat cttcaaacct ttcgtccgat ccatcttgca ggctacctct 2941 cgaacgaact atcgcaagtc tcttggccgg ccttgcgcct tggctattgc ttggcagcgc 3001 ctatcgccag gtattactcc aatcccgaat atccgagatc gggatcaccc gagagaagtt 3061 caacctacat cctcaatccc gatctatccg agatccgagg aatatcgaaa tcggggcgcg 3121 cctggcctcc gcgccgggtt ttggcgcctc ccgcgggcgc ccccctcgtc acggcgagcg 3181 ctgccacgtc agacgaaggg cgcaggagcg tcctgatcct tccgcccgga cgctcaggac 3241 agcggcccgc tgctcataag actcggcctt agaaccccag tatcagcaga aggacatttt 3301 aggacgggac ttgggtgact ctagggcact ggttttcttt ccagagagcg gaacaggcga 3361 ggaaaagtag tcccttctcg gcgattctgc ggagggatct ccgtggggcg gtgaacgccg 3421 atgattatat aaggacgcgc cgggtgtggc acagctagtt ccgtcgcagc cgggatttgg 3481 gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg gtgagtagcg ggctgctggg 3541 ctggccgggg ctttcgtggc cgccgggccg ctcggtggga cggaagcgtg tggagagacc 3601 gccaagggct gtagtctggg tccgcgagca aggttgccct gaactggggg ttggggggag 3661 cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag acgcttgtga ggcgggctgt 3721 gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa gaacccaagg tcttgagccc 3781 ttcgctaatg cgggaaagct cttattcggg tgagatgggc tgggcaccat ctggggaccc 3841 tgacgtgaag tttgtcactg actggagaac tcggtttgtc gtctgttgcg ggggcggcag 3901 ttatggcggt gccgttgggc agtgcacccg tacctttggg agcgcgcgcc ctcgtcgtgt 3961 cgtgacgtca cccgttctgt tggcttataa tgcagggtgg ggccacctgc cggtaggtgt 4021 gcggtaggct tttctccgtc gcaggacgca gggttcgggc ctagggtagg ctctcctgaa 4081 tcgacaggcg ccggacctct ggtgagggga gggataagtg aggcgtcagt ttctttggtc 4141 ggttttatgt acctatcttc ttaagtagct gaagctccgg ttttgaacta tgcgctcggg 4201 gttggcgagt gtgttttgtg aagtttttta ggcacctttt gaaatgtaat catttgggtc 4261 aatatgtaat tttcagtgtt agacttgtaa attgtccgct aaattctggc cgtttttggc 4321 ttttttgtta gacaacatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga 4381 cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca 4441 caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac 4501 gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt 4561 ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat 4621 ggccgagttg agcggttccc ggctggccgc gcagcaacag atggagggcc tcctggcgcc 4681 gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca 4741 gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggccgccg agcgcgccgg 4801 ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc cccttctacg agcggctcgg 4861 cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg 4921 caagcccggt gcctaggcta gtatgtaagc ctagtcttag ataataaaat cgctatccat 4981 cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct ggtgtaccga 5041 gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag tcagccagtc 5101 ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa gcctggtcac 5161 ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca acgtataatc 5221 gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc tttcgcatga 5281 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt 5341 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcgcgag 5401 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag 5461 aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta 5521 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg 5581 agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca 5641 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgattggag 5701 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc 5761 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg 5821 tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact ctagcttccc 5881 ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg 5941 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg 6001 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga 6061 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac 6121 tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg cctgatgcga 6181 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac 6241 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc ccgaatcgtt 6301 taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga acagccgtgg 6361 cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac ctttttggca 6421 gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct tccctctagt 6481 ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga cccgtttaga 6541 ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc aacacacatc 6601 catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc cagatctagt 6661 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta 6721 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga 6781 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt 6841 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta 6901 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg 6961 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg //