LOCUS Exported 7418 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 N (nucleocapsid). ACCESSION . VERSION . KEYWORDS pGBW-m4133969 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7418) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7418) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7418 /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..1327 /codon_start=1 /product="affinity and epitope tag derived from pancreatic ribonuclease A" /label=S-Tag /translation="KETAAAKFERQHMDS" polyA_signal 1380..1501 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1391..1410) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1445..1464 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1616..1662 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1692..1828 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1873..1944 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2170..2758) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2250..2269) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 3258..4467 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 4304..4323 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 4471..5070 /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(4471..4490) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4967..4987 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 5097..5145 /note="synthetic polyadenylation signal" CDS 5411..6271 /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(5629..5648) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 6284..6420 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6572..6643 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6648..6694 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6712..6760 /note="synthetic polyadenylation signal" enhancer 6841..7144 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 7146..7348 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 7299..7319 /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 cgaaagaaac cgcggcagct aagttcgaga ggcagcacat 1321 ggacagttaa gctagcttga ctgactgaga tacagcgtac cttcagctca cagacatgat 1381 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat 1441 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt 1501 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt 1561 ttaaagcaag taaaacctct acaaatgtgg tattggccca tctctatcgg tatcgtagca 1621 taaccccttg gggcctctaa acgggtcttg aggggttttt tgtgcccctc gggccggatt 1681 gctatctacc ggcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact 1741 cccacatatg ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt 1801 gtcctcctta ccagaaattt atccttaagg tcgtcagcta tcctgcaggc gatctctcga 1861 tttcgatcaa gacattcctt taatggtctt ttctggacac cactaggggt cagaagtagt 1921 tcatcaaact ttcttccctc cctaatctca ttggttacct tgggctatcg aaacttaatt 1981 aaccagtcaa gtcagctact tggcgagatc gacttgtctg ggtttcgact acgctcagaa 2041 ttgcgtcagt caagttcgat ctggtccttg ctattgcacc cgttctccga ttacgagttt 2101 catttaaatc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 2161 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 2221 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 2281 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 2341 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 2401 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 2461 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 2521 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 2581 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 2641 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 2701 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 2761 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 2821 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 2881 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 2941 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcatttaa 3001 atttccgaac tctccaaggc cctcgtcgga aaatcttcaa acctttcgtc cgatccatct 3061 tgcaggctac ctctcgaacg aactatcgca agtctcttgg ccggccttgc gccttggcta 3121 ttgcttggca gcgcctatcg ccaggtatta ctccaatccc gaatatccga gatcgggatc 3181 acccgagaga agttcaacct acatcctcaa tcccgatcta tccgagatcc gaggaatatc 3241 gaaatcgggg cgcgcctggc ctccgcgccg ggttttggcg cctcccgcgg gcgcccccct 3301 cgtcacggcg agcgctgcca cgtcagacga agggcgcagg agcgtcctga tccttccgcc 3361 cggacgctca ggacagcggc ccgctgctca taagactcgg ccttagaacc ccagtatcag 3421 cagaaggaca ttttaggacg ggacttgggt gactctaggg cactggtttt ctttccagag 3481 agcggaacag gcgaggaaaa gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg 3541 ggcggtgaac gccgatgatt atataaggac gcgccgggtg tggcacagct agttccgtcg 3601 cagccgggat ttgggtcgcg gttcttgttt gtggatcgct gtgatcgtca cttggtgagt 3661 agcgggctgc tgggctggcc ggggctttcg tggccgccgg gccgctcggt gggacggaag 3721 cgtgtggaga gaccgccaag ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg 3781 ggggttgggg ggagcgcagc aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt 3841 gtgaggcggg ctgtgaggtc gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc 3901 aaggtcttga gcccttcgct aatgcgggaa agctcttatt cgggtgagat gggctgggca 3961 ccatctgggg accctgacgt gaagtttgtc actgactgga gaactcggtt tgtcgtctgt 4021 tgcgggggcg gcagttatgg cggtgccgtt gggcagtgca cccgtacctt tgggagcgcg 4081 cgccctcgtc gtgtcgtgac gtcacccgtt ctgttggctt ataatgcagg gtggggccac 4141 ctgccggtag gtgtgcggta ggcttttctc cgtcgcagga cgcagggttc gggcctaggg 4201 taggctctcc tgaatcgaca ggcgccggac ctctggtgag gggagggata agtgaggcgt 4261 cagtttcttt ggtcggtttt atgtacctat cttcttaagt agctgaagct ccggttttga 4321 actatgcgct cggggttggc gagtgtgttt tgtgaagttt tttaggcacc ttttgaaatg 4381 taatcatttg ggtcaatatg taattttcag tgttagactt gtaaattgtc cgctaaattc 4441 tggccgtttt tggctttttt gttagacaac atgaccgagt acaagcccac ggtgcgcctc 4501 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac 4561 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa 4621 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc 4681 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag 4741 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggag 4801 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc 4861 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcc 4921 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc 4981 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc 5041 tggtgcatga cccgcaagcc cggtgcctag gctagtatgt aagcctagtc ttagataata 5101 aaatcgctat ccatcgaaga tggatgtgtg ttggtttttt gtgtgtgtaa cgctaggcgc 5161 gcctggtgta ccgagaacga tcctctcagt gcgagtctcg acgatccata tcgttgcttg 5221 gcagtcagcc agtcggaatc cagcttggga cccaggaagt ccaatcgtca gatattgtac 5281 tcaagcctgg tcacggcagc gtaccgatct gtttaaacct agatattgat agtctgatcg 5341 gtcaacgtat aatcgagtcc tagcttttgc aaacatctat caagagacag gatcagcagg 5401 aggctttcgc atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt 5461 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca 5521 gttgggtgcg cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag 5581 ttttcgcccc gaagaacgct ttccaatgat gagcactttt aaagttctgc tatgtggcgc 5641 ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca 5701 gaatgacttg gttgagtatt caccagtcac agaaaagcat cttacggatg gcatgacagt 5761 aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct 5821 gacaacgatt ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt 5881 aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga 5941 caccacgatg cctgtagcaa tggcaacaac cttgcgtaaa ctattaactg gcgaactact 6001 tactctagct tcccggcaac agttgataga ctggatggag gcggataaag ttgcaggacc 6061 acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga 6121 gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt 6181 agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga 6241 gataggtgcc tcactgatta agcattggta accgattcta ggtgcattgg cgcagaaaaa 6301 aatgcctgat gcgacgctgc gcgtcttata ctcccacata tgccagattc agcaacggat 6361 acggcttccc caacttgccc acttccatac gtgtcctcct taccagaaat ttatccttaa 6421 gatcccgaat cgtttaaact cgactctggc tctatcgaat ctccgtcgtt tcgagcttac 6481 gcgaacagcc gtggcgctca tttgctcgtc gggcatcgaa tctcgtcagc tatcgtcagc 6541 ttaccttttt ggcagcgatc gcggctcccg acatcttgga ccattagctc cacaggtatc 6601 ttcttccctc tagtggtcat aacagcagct tcagctacct ctcaattcaa aaaacccctc 6661 aagacccgtt tagaggcccc aaggggttat gctatcaatc gttgcgttac acacacaaaa 6721 aaccaacaca catccatctt cgatggatag cgattttatt atctaactgc tgatcgagtg 6781 tagccagatc tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg 6841 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 6901 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 6961 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 7021 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 7081 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 7141 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 7201 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 7261 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 7321 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc 7381 gatcctacca tccactcgac acacccgcca gcggccgc //