LOCUS Exported 6395 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp7. ACCESSION . VERSION . KEYWORDS pGBW-m4252809 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6395) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6395) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6395 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 257..277 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 278..304 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" polyA_signal 357..478 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(368..387) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 422..441 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 593..639 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 669..805 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 850..921 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1147..1735) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1227..1246) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 2235..3444 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3281..3300 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 3448..4047 /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(3448..3467) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 3944..3964 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 4074..4122 /note="synthetic polyadenylation signal" CDS 4388..5248 /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(4606..4625) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5261..5397 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 5549..5620 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 5625..5671 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5689..5737 /note="synthetic polyadenylation signal" enhancer 5818..6121 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 6123..6325 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6276..6296 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgagc aaaatgtctg acgtcaagtg tacatccgta gttctgttat cagtgcttca 61 acagctgcgg gtggagagtt cgtcaaagct ctgggcccag tgcgtgcagc tgcacaatga 121 tatattgctc gcgaaagata ctacggaagc tttcgagaag atggtcagct tgctatctgt 181 tctactgagc atgcaaggag cagtagacat caacaaactt tgcgaagaga tgttagacaa 241 ccgagccacc ctgcaagaaa acctttactt tcagtcgtac ccctatgacg taccagatta 301 tgcgtaagct agcttgactg actgagatac agcgtacctt cagctcacag acatgataag 361 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg 421 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa 481 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta 541 aagcaagtaa aacctctaca aatgtggtat tggcccatct ctatcggtat cgtagcataa 601 ccccttgggg cctctaaacg ggtcttgagg ggttttttgt gcccctcggg ccggattgct 661 atctaccggc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc 721 acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc catacgtgtc 781 ctccttacca gaaatttatc cttaaggtcg tcagctatcc tgcaggcgat ctctcgattt 841 cgatcaagac attcctttaa tggtcttttc tggacaccac taggggtcag aagtagttca 901 tcaaactttc ttccctccct aatctcattg gttaccttgg gctatcgaaa cttaattaac 961 cagtcaagtc agctacttgg cgagatcgac ttgtctgggt ttcgactacg ctcagaattg 1021 cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt tctccgatta cgagtttcat 1081 ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct 1141 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca 1201 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct 1261 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc 1321 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt 1381 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc 1441 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc 1501 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg 1561 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc 1621 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag 1681 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga 1741 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat 1801 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag 1861 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat 1921 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg catttaaatt 1981 tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc tttcgtccga tccatcttgc 2041 aggctacctc tcgaacgaac tatcgcaagt ctcttggccg gccttgcgcc ttggctattg 2101 cttggcagcg cctatcgcca ggtattactc caatcccgaa tatccgagat cgggatcacc 2161 cgagagaagt tcaacctaca tcctcaatcc cgatctatcc gagatccgag gaatatcgaa 2221 atcggggcgc gcctggcctc cgcgccgggt tttggcgcct cccgcgggcg cccccctcgt 2281 cacggcgagc gctgccacgt cagacgaagg gcgcaggagc gtcctgatcc ttccgcccgg 2341 acgctcagga cagcggcccg ctgctcataa gactcggcct tagaacccca gtatcagcag 2401 aaggacattt taggacggga cttgggtgac tctagggcac tggttttctt tccagagagc 2461 ggaacaggcg aggaaaagta gtcccttctc ggcgattctg cggagggatc tccgtggggc 2521 ggtgaacgcc gatgattata taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag 2581 ccgggatttg ggtcgcggtt cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc 2641 gggctgctgg gctggccggg gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt 2701 gtggagagac cgccaagggc tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg 2761 gttgggggga gcgcagcaaa atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg 2821 aggcgggctg tgaggtcgtt gaaacaaggt ggggggcatg gtgggcggca agaacccaag 2881 gtcttgagcc cttcgctaat gcgggaaagc tcttattcgg gtgagatggg ctgggcacca 2941 tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt cgtctgttgc 3001 gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc 3061 cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg gggccaccag 3121 ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg cctagggtag 3181 gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt gaggcgtcag 3241 tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg gttttgaact 3301 atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa 3361 tcatttgggt caatatgtaa ttttcagtgt tagacttgta aattgtccgc taaattctgg 3421 ccgtttttgg cttttttgtt agacaacatg accgagtaca agcccacggt gcgcctcgcc 3481 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc 3541 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa 3601 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc 3661 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc 3721 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggagggc 3781 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg 3841 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggccgcc 3901 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac 3961 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg 4021 tgcatgaccc gcaagcccgg tgcctaggct agtatgtaag cctagtctta gataataaaa 4081 tcgctatcca tcgaagatgg atgtgtgttg gttttttgtg tgtgtaacgc taggcgcgcc 4141 tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg ttgcttggca 4201 gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat attgtactca 4261 agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt ctgatcggtc 4321 aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat cagcaggagg 4381 ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg 4441 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt 4501 gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt 4561 tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat gtggcgcggt 4621 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa 4681 tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca tgacagtaag 4741 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac 4801 aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac 4861 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac 4921 cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg aactacttac 4981 tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg caggaccact 5041 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg 5101 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt 5161 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat 5221 aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc agaaaaaaat 5281 gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc aacggatacg 5341 gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta tccttaagat 5401 cccgaatcgt ttaaactcga ctctggctct atcgaatctc cgtcgtttcg agcttacgcg 5461 aacagccgtg gcgctcattt gctcgtcggg catcgaatct cgtcagctat cgtcagctta 5521 cctttttggc agcgatcgcg gctcccgaca tcttggacca ttagctccac aggtatcttc 5581 ttccctctag tggtcataac agcagcttca gctacctctc aattcaaaaa acccctcaag 5641 acccgtttag aggccccaag gggttatgct atcaatcgtt gcgttacaca cacaaaaaac 5701 caacacacat ccatcttcga tggatagcga ttttattatc taactgctga tcgagtgtag 5761 ccagatctag taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt 5821 tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac 5881 gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg 5941 ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag 6001 tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat 6061 gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat 6121 gctgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt 6181 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga 6241 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg 6301 gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatcagat ctttgtcgat 6361 cctaccatcc actcgacaca cccgccagcg gccgc //