LOCUS Exported 6171 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp2. ACCESSION . VERSION . KEYWORDS pGBW-m4134181 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6171) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6171) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6171 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 1922..1942 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1943..1987 /codon_start=1 /product="affinity and epitope tag derived from pancreatic ribonuclease A" /label=S-Tag /translation="KETAAAKFERQHMDS" polyA_signal 2040..2161 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2051..2070) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2105..2124 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2276..2322 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2352..2488 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2533..2604 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2830..3418) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2910..2929) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 4164..5024 /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(4382..4401) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5037..5173 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 5325..5396 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 5401..5447 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5465..5513 /note="synthetic polyadenylation signal" enhancer 5594..5897 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 5899..6101 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6052..6072 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 6170..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggcc tacacaagat atgtggataa caatttttgt gggccggacg gatacccttt 61 agaatgcatc aaagatttgc ttgcaagggc gggcaaggct tcgtgcactc tgtccgagca 121 gctggacttc attgatacca agcgcggtgt ttattgttgc cgagagcatg aacacgagat 181 agcctggtat acggaacggt cagagaaaag ttacgaactc caaaccccat tcgagatcaa 241 gctagctaaa aagtttgaca ctttcaacgg cgaatgtccc aattttgtct ttcccctgaa 301 ctctattatc aaaacaatac agcctcgggt ggagaagaag aaattagacg gattcatggg 361 gcgtattagg agcgtatacc cagtggccag cccgaatgaa tgcaaccaga tgtgtctttc 421 cacactgatg aagtgcgatc actgtggcga gacttcttgg caaaccggtg acttcgtaaa 481 agcgacatgc gagttttgtg gtaccgagaa tctgactaag gagggagcta ccacatgcgg 541 ctatttgcca cagaatgcag tggtcaagat ctactgtcct gcatgccata actctgaagt 601 cggccccgag cactcgctcg ccgaatatca caatgagagc gggctaaaaa ctatactgag 661 gaaaggaggt cgcaccatcg cctttggggg ctgcgttttc tcctacgtgg ggtgtcataa 721 caaatgtgcc tattgggttc ccagagcctc cgctaacatt gggtgcaatc atacgggagt 781 ggtcggggaa ggctcagaag gcttgaacga taacctcctt gaaattctgc aaaaggagaa 841 ggtgaatatc aatatcgttg gcgacttcaa gttaaatgag gaaatcgcga ttattctggc 901 aagtttttct gcctcaacta gcgcctttgt agagacagtg aaaggactcg attataaagc 961 tttcaaacag atagtcgaga gttgtggtaa cttcaaggtt acaaagggca aggctaaaaa 1021 gggtgcttgg aacattggag agcagaagtc aatcttgagt ccactgtacg cgtttgcatc 1081 tgaggccgcc cgtgtggtca ggagcatatt cagtcgaacc ctagaaaccg cacagaatag 1141 cgtgcgggta ttacagaagg cggctattac tattctcgac ggtatctcgc agtattctct 1201 gagactgatc gacgccatga tgtttacaag cgatctggca acgaataact tagtggtcat 1261 ggcgtacata accggagggg tcgtgcaact tacctcccag tggctaacga acatctttgg 1321 gacggtttac gagaagctaa aacctgtgct ggactggctc gaagagaaat tcaaagaagg 1381 cgtggaattt ttgcgtgacg ggtgggagat cgtaaagttc atatcaacat gtgcttgcga 1441 aatcgttggc ggacaaatcg taacttgcgc aaaagagatt aaagaatcgg tgcagacctt 1501 ctttaaactt gttaacaaat tcctggcttt gtgtgccgat tctattatca tcgggggagc 1561 aaagcttaaa gccctaaatt taggcgagac atttgtgacg cacagtaagg gcctgtatag 1621 aaagtgcgtg aagagtcgcg aagaaaccgg actgctcatg cccttaaaag cgccaaaaga 1681 aataattttc ctggaagggg agacattgcc cactgaagtg ctcactgagg aagtagttct 1741 gaaaactggg gatctgcaac ctttagagca gccgacctcc gaggccgtcg aggcacctct 1801 cgtgggaacc ccggtctgta ttaatgggtt gatgctgctc gagattaagg ataccgaaaa 1861 atactgcgca ctagccccca acatgatggt gactaacaac acatttaccc tcaagggagg 1921 ggaaaacctt tactttcagt cgaaagaaac cgcggcagct aagttcgaga ggcagcacat 1981 ggacagttaa gctagcttga ctgactgaga tacagcgtac cttcagctca cagacatgat 2041 aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa aatgctttat 2101 ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca ataaacaagt 2161 taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt gggaggtttt 2221 ttaaagcaag taaaacctct acaaatgtgg tattggccca tctctatcgg tatcgtagca 2281 taaccccttg gggcctctaa acgggtcttg aggggttttt tgtgcccctc gggccggatt 2341 gctatctacc ggcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact 2401 cccacatatg ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt 2461 gtcctcctta ccagaaattt atccttaagg tcgtcagcta tcctgcaggc gatctctcga 2521 tttcgatcaa gacattcctt taatggtctt ttctggacac cactaggggt cagaagtagt 2581 tcatcaaact ttcttccctc cctaatctca ttggttacct tgggctatcg aaacttaatt 2641 aaccagtcaa gtcagctact tggcgagatc gacttgtctg ggtttcgact acgctcagaa 2701 ttgcgtcagt caagttcgat ctggtccttg ctattgcacc cgttctccga ttacgagttt 2761 catttaaatc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt 2821 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag 2881 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc 2941 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc 3001 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt 3061 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt 3121 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc 3181 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa 3241 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa 3301 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg 3361 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga 3421 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg 3481 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg 3541 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt 3601 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcatttaa 3661 atttccgaac tctccaaggc cctcgtcgga aaatcttcaa acctttcgtc cgatccatct 3721 tgcaggctac ctctcgaacg aactatcgca agtctcttgg ccggccttgc gccttggcta 3781 ttgcttggca gcgcctatcg ccaggtatta ctccaatccc gaatatccga gatcgggatc 3841 acccgagaga agttcaacct acatcctcaa tcccgatcta tccgagatcc gaggaatatc 3901 gaaatcgggg cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc 3961 atatcgttgc ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg 4021 tcagatattg tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt 4081 gatagtctga tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga 4141 caggatcagc aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct 4201 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag 4261 atgctgaaga tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta 4321 agatccttga gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc 4381 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca 4441 tacactattc tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg 4501 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg 4561 ccaacttact tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca 4621 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa 4681 acgacgagcg tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa 4741 ctggcgaact acttactcta gcttcccggc aacagttgat agactggatg gaggcggata 4801 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat 4861 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc 4921 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata 4981 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat 5041 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga 5101 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga 5161 aatttatcct taagatcccg aatcgtttaa actcgactct ggctctatcg aatctccgtc 5221 gtttcgagct tacgcgaaca gccgtggcgc tcatttgctc gtcgggcatc gaatctcgtc 5281 agctatcgtc agcttacctt tttggcagcg atcgcggctc ccgacatctt ggaccattag 5341 ctccacaggt atcttcttcc ctctagtggt cataacagca gcttcagcta cctctcaatt 5401 caaaaaaccc ctcaagaccc gtttagaggc cccaaggggt tatgctatca atcgttgcgt 5461 tacacacaca aaaaaccaac acacatccat cttcgatgga tagcgatttt attatctaac 5521 tgctgatcga gtgtagccag atctagtaat caattacggg gtcattagtt catagcccat 5581 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 5641 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 5701 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 5761 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 5821 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 5881 tcatcgctat taccatgctg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 5941 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 6001 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 6061 gcggtaggcg tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga 6121 tcagatcttt gtcgatccta ccatccactc gacacacccg ccagcggccg c //