LOCUS Exported 4530 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp7. ACCESSION . VERSION . KEYWORDS pGBW-m4134071 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4530) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 4530) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4530 /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 281..346 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" polyA_signal 399..520 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(410..429) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 464..483 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 635..681 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 711..847 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 892..963 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1189..1777) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1269..1288) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 2523..3383 /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(2741..2760) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 3396..3532 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3684..3755 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 3760..3806 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 3824..3872 /note="synthetic polyadenylation signal" enhancer 3953..4256 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4258..4460 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4411..4431 /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 tcagtcgatg gattataagg accacgatgg 301 agactacaaa gatcatgaca ttgattacaa ggacgacgat gacaaataag ctagcttgac 361 tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg atgagtttgg 421 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat 481 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca 541 ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt aaaacctcta 601 caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg ggcctctaaa 661 cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg gcattggcgc 721 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc 781 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta 841 tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag acattccttt 901 aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt tcttccctcc 961 ctaatctcat tggttacctt gggctatcga aacttaatta accagtcaag tcagctactt 1021 ggcgagatcg acttgtctgg gtttcgacta cgctcagaat tgcgtcagtc aagttcgatc 1081 tggtccttgc tattgcaccc gttctccgat tacgagtttc atttaaatca tgtgagcaaa 1141 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct 1201 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac 1261 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc 1321 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc 1381 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg 1441 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga 1501 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag 1561 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta 1621 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag 1681 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg 1741 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac 1801 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc 1861 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag 1921 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc 1981 agcgatctgt ctatttcgtt catccatagt tgcatttaaa tttccgaact ctccaaggcc 2041 ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt gcaggctacc tctcgaacga 2101 actatcgcaa gtctcttggc cggccttgcg ccttggctat tgcttggcag cgcctatcgc 2161 caggtattac tccaatcccg aatatccgag atcgggatca cccgagagaa gttcaaccta 2221 catcctcaat cccgatctat ccgagatccg aggaatatcg aaatcggggc gcgcctggtg 2281 taccgagaac gatcctctca gtgcgagtct cgacgatcca tatcgttgct tggcagtcag 2341 ccagtcggaa tccagcttgg gacccaggaa gtccaatcgt cagatattgt actcaagcct 2401 ggtcacggca gcgtaccgat ctgtttaaac ctagatattg atagtctgat cggtcaacgt 2461 ataatcgagt cctagctttt gcaaacatct atcaagagac aggatcagca ggaggctttc 2521 gcatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc 2581 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg 2641 cgcgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc 2701 ccgaagaacg ctttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat 2761 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact 2821 tggttgagta ttcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat 2881 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga 2941 ttggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc 3001 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga 3061 tgcctgtagc aatggcaaca accttgcgta aactattaac tggcgaacta cttactctag 3121 cttcccggca acagttgata gactggatgg aggcggataa agttgcagga ccacttctgc 3181 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt 3241 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct 3301 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg 3361 cctcactgat taagcattgg taaccgattc taggtgcatt ggcgcagaaa aaaatgcctg 3421 atgcgacgct gcgcgtctta tactcccaca tatgccagat tcagcaacgg atacggcttc 3481 cccaacttgc ccacttccat acgtgtcctc cttaccagaa atttatcctt aagatcccga 3541 atcgtttaaa ctcgactctg gctctatcga atctccgtcg tttcgagctt acgcgaacag 3601 ccgtggcgct catttgctcg tcgggcatcg aatctcgtca gctatcgtca gcttaccttt 3661 ttggcagcga tcgcggctcc cgacatcttg gaccattagc tccacaggta tcttcttccc 3721 tctagtggtc ataacagcag cttcagctac ctctcaattc aaaaaacccc tcaagacccg 3781 tttagaggcc ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa aaaaccaaca 3841 cacatccatc ttcgatggat agcgatttta ttatctaact gctgatcgag tgtagccaga 3901 tctagtaatc aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat 3961 aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa 4021 taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg 4081 agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc 4141 cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct 4201 tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt accatgctga 4261 tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa 4321 gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc 4381 caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg 4441 aggtctatat aagcagagct ggtttagtga accgtcagat cagatctttg tcgatcctac 4501 catccactcg acacacccgc cagcggccgc //