LOCUS Exported 7408 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 N (nucleocapsid). ACCESSION . VERSION . KEYWORDS pGBW-m4133222 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7408) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7408) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7408 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 1262..1282 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1283..1324 /codon_start=1 /product="epitope tag from simian virus 5" /label=V5 tag /translation="GKPIPNPLLGLDST" polyA_signal 1377..1498 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1388..1407) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1442..1461 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1613..1659 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1689..1825 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1870..1941 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2167..2755) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2247..2266) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 3255..4464 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 4301..4320 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 4468..5067 /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(4468..4487) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4964..4984 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 5094..5142 /note="synthetic polyadenylation signal" CDS 5408..6268 /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(5626..5645) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 6281..6417 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6562..6633 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6638..6684 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6702..6750 /note="synthetic polyadenylation signal" enhancer 6831..7134 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 7136..7338 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 7289..7309 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgtct gataatggac cccaaaatca gcgaaatgca ccccgcatta cgtttggtgg 61 accctcagat tcaactggca gtaaccagaa tggagaacgc agtggggcgc gatcaaaaca 121 acgtcggccc caaggtttac ccaataatac tgcgtcttgg ttcaccgctc tcactcaaca 181 tggcaaggaa gaccttaaat tccctcgagg acaaggcgtt ccaattaaca ccaatagcag 241 tccagatgac caaattggct actaccgaag agctaccaga cgaattcgtg gtggtgacgg 301 taaaatgaaa gatctcagtc caagatggta tttctactac ctaggaactg ggccagaagc 361 tggacttccc tatggtgcta acaaagacgg catcatatgg gttgcaactg agggagcctt 421 gaatacacca aaagatcaca ttggcacccg caatcctgct aacaatgctg caatcgtgct 481 acaacttcct caaggaacaa cattgccaaa aggcttctac gcagaaggga gcagaggcgg 541 cagtcaagcc tcttctcgtt cctcatcacg tagtcgcaac agttcaagaa attcaactcc 601 aggcagcagt aggggaactt ctcctgctag aatggctggc aatggcggtg atgctgctct 661 tgctttgctg ctgcttgaca gattgaacca gcttgagagc aaaatgtctg gtaaaggcca 721 acaacaacaa ggccaaactg tcactaagaa atctgctgct gaggcttcta agaagcctcg 781 gcaaaaacgt actgccacta aagcatacaa tgtaacacaa gctttcggca gacgtggtcc 841 agaacaaacc caaggaaatt ttggggacca ggaactaatc agacaaggaa ctgattacaa 901 acattggccg caaattgcac aatttgcccc cagcgcttca gcgttcttcg gaatgtcgcg 961 cattggcatg gaagtcacac cttcgggaac gtggttgacc tacacaggtg ccatcaaatt 1021 ggatgacaaa gatccaaatt tcaaagatca agtcattttg ctgaataagc atattgacgc 1081 atacaaaaca ttcccaccaa cagagcctaa aaaggacaaa aagaagaagg ctgatgaaac 1141 tcaagcctta ccgcagagac agaagaaaca gcaaactgtg actcttcttc ctgctgcaga 1201 tttggatgat ttctccaaac aattgcaaca atccatgagc agtgctgact caactcaggc 1261 cgaaaacctt tactttcagt cggggaaacc tatacccaat ccacttctcg gcttagattc 1321 tacctaagct agcttgactg actgagatac agcgtacctt cagctcacag acatgataag 1381 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg 1441 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa 1501 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta 1561 aagcaagtaa aacctctaca aatgtggtat tggcccatct ctatcggtat cgtagcataa 1621 ccccttgggg cctctaaacg ggtcttgagg ggttttttgt gcccctcggg ccggattgct 1681 atctaccggc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc 1741 acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc catacgtgtc 1801 ctccttacca gaaatttatc cttaaggtcg tcagctatcc tgcaggcgat ctctcgattt 1861 cgatcaagac attcctttaa tggtcttttc tggacaccac taggggtcag aagtagttca 1921 tcaaactttc ttccctccct aatctcattg gttaccttgg gctatcgaaa cttaattaac 1981 cagtcaagtc agctacttgg cgagatcgac ttgtctgggt ttcgactacg ctcagaattg 2041 cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt tctccgatta cgagtttcat 2101 ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct 2161 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca 2221 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct 2281 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc 2341 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt 2401 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc 2461 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc 2521 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg 2581 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc 2641 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag 2701 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga 2761 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat 2821 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag 2881 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat 2941 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg catttaaatt 3001 tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc tttcgtccga tccatcttgc 3061 aggctacctc tcgaacgaac tatcgcaagt ctcttggccg gccttgcgcc ttggctattg 3121 cttggcagcg cctatcgcca ggtattactc caatcccgaa tatccgagat cgggatcacc 3181 cgagagaagt tcaacctaca tcctcaatcc cgatctatcc gagatccgag gaatatcgaa 3241 atcggggcgc gcctggcctc cgcgccgggt tttggcgcct cccgcgggcg cccccctcgt 3301 cacggcgagc gctgccacgt cagacgaagg gcgcaggagc gtcctgatcc ttccgcccgg 3361 acgctcagga cagcggcccg ctgctcataa gactcggcct tagaacccca gtatcagcag 3421 aaggacattt taggacggga cttgggtgac tctagggcac tggttttctt tccagagagc 3481 ggaacaggcg aggaaaagta gtcccttctc ggcgattctg cggagggatc tccgtggggc 3541 ggtgaacgcc gatgattata taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag 3601 ccgggatttg ggtcgcggtt cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc 3661 gggctgctgg gctggccggg gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt 3721 gtggagagac cgccaagggc tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg 3781 gttgggggga gcgcagcaaa atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg 3841 aggcgggctg tgaggtcgtt gaaacaaggt ggggggcatg gtgggcggca agaacccaag 3901 gtcttgagcc cttcgctaat gcgggaaagc tcttattcgg gtgagatggg ctgggcacca 3961 tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt cgtctgttgc 4021 gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc 4081 cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg gggccacctg 4141 ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg cctagggtag 4201 gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt gaggcgtcag 4261 tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg gttttgaact 4321 atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa 4381 tcatttgggt caatatgtaa ttttcagtgt tagacttgta aattgtccgc taaattctgg 4441 ccgtttttgg cttttttgtt agacaacatg accgagtaca agcccacggt gcgcctcgcc 4501 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc 4561 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa 4621 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc 4681 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc 4741 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggagggc 4801 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg 4861 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggccgcc 4921 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac 4981 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg 5041 tgcatgaccc gcaagcccgg tgcctaggct agtatgtaag cctagtctta gataataaaa 5101 tcgctatcca tcgaagatgg atgtgtgttg gttttttgtg tgtgtaacgc taggcgcgcc 5161 tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg ttgcttggca 5221 gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat attgtactca 5281 agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt ctgatcggtc 5341 aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat cagcaggagg 5401 ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg 5461 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt 5521 gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt 5581 tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat gtggcgcggt 5641 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa 5701 tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca tgacagtaag 5761 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac 5821 aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac 5881 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac 5941 cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg aactacttac 6001 tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg caggaccact 6061 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg 6121 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt 6181 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat 6241 aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc agaaaaaaat 6301 gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc aacggatacg 6361 gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta tccttaagat 6421 cgtttaaact cgactctggc tctatcgaat ctccgtcgtt tcgagcttac gcgaacagcc 6481 gtggcgctca tttgctcgtc gggcatcgaa tctcgtcagc tatcgtcagc ttaccttttt 6541 ggcagcgatc gcggctcccg acatcttgga ccattagctc cacaggtatc ttcttccctc 6601 tagtggtcat aacagcagct tcagctacct ctcaattcaa aaaacccctc aagacccgtt 6661 tagaggcccc aaggggttat gctatcaatc gttgcgttac acacacaaaa aaccaacaca 6721 catccatctt cgatggatag cgattttatt atctaactgc tgatcgagtg tagccagatc 6781 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa 6841 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata 6901 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag 6961 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc 7021 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta 7081 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catgctgatg 7141 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt 7201 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca 7261 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag 7321 gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc gatcctacca 7381 tccactcgac acacccgcca gcggccgc //