LOCUS Exported 7020 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 RNA-dependent RNA polymerase. ACCESSION . VERSION . KEYWORDS pGBW-m4133484 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7020) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7020) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7020 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 2798..2818 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 2819..2836 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 2889..3010 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2900..2919) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2954..2973 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 3125..3171 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 3201..3337 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3382..3453 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(3679..4267) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3759..3778) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 5013..5873 /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(5231..5250) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5886..6022 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6174..6245 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6250..6296 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6314..6362 /note="synthetic polyadenylation signal" enhancer 6443..6746 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 6748..6950 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6901..6921 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 7019..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggct gatgcacaat cgtttttaaa ccgggtttgc ggtgtaagtg cagcccgtct 61 tacaccgtgc ggcacaggca ctagtactga tgtcgtatac agggcttttg acatctacaa 121 tgataaagta gctggttttg ctaaattcct aaaaactaat tgttgtcgct tccaagaaaa 181 ggacgaagat gacaatttaa ttgattctta ctttgtagtt aagagacaca ctttctctaa 241 ctaccaacat gaagaaacaa tttataattt acttaaggat tgtccagctg ttgctaaaca 301 tgacttcttt aagtttagaa tagacggtga catggtacca catatatcac gtcaacgtct 361 tactaaatac acaatggcag acctcgtcta tgctttaagg cattttgatg aaggtaattg 421 tgacacatta aaagaaatac ttgtcacata caattgttgt gatgatgatt atttcaataa 481 aaaggactgg tatgattttg tagaaaaccc agatatatta cgcgtatacg ccaacttagg 541 tgaacgtgta cgccaagctt tgttaaaaac agtacaattc tgtgatgcca tgcgaaatgc 601 tggtattgtt ggtgtactga cattagataa tcaagatctc aatggtaact ggtatgattt 661 cggtgatttc atacaaacca cgccaggtag tggagttcct gttgtagatt cttattattc 721 attgttaatg cctatattaa ccttgaccag ggctttaact gcagagtcac atgttgacac 781 tgacttaaca aagccttaca ttaagtggga tttgttaaaa tatgacttca cggaagagag 841 gttaaaactc tttgaccgtt attttaaata ttgggatcag acataccacc caaattgtgt 901 taactgtttg gatgacagat gcattctgca ttgtgcaaac tttaatgttt tattctctac 961 agtgttccca cctacaagtt ttggaccact agtgagaaaa atatttgttg atggtgttcc 1021 atttgtagtt tcaactggat accacttcag agagctaggt gttgtacata atcaggatgt 1081 aaacttacat agctctagac ttagttttaa ggaattactt gtgtatgctg ctgaccctgc 1141 tatgcacgct gcttctggta atctattact agataaacgc actacgtgct tttcagtagc 1201 tgcacttact aacaatgttg cttttcaaac tgtcaaaccc ggtaatttta acaaagactt 1261 ctatgacttt gctgtgtcta agggtttctt taaggaagga agttctgttg aattaaaaca 1321 cttcttcttt gctcaggatg gtaatgctgc tatcagcgat tatgactact atcgttataa 1381 tctaccaaca atgtgtgata tcagacaact actatttgta gttgaagttg ttgataagta 1441 ctttgattgt tacgatggtg gctgtattaa tgctaaccaa gtcatcgtca acaacctaga 1501 caaatcagct ggttttccat ttaataaatg gggtaaggct agactttatt atgattcaat 1561 gagttatgag gatcaagatg cacttttcgc atatacaaaa cgtaatgtca tccctactat 1621 aactcaaatg aatcttaagt atgccattag tgcaaagaat agagctcgca ccgtagctgg 1681 tgtctctatc tgtagtacta tgaccaatag acagtttcat caaaaattat tgaaatcaat 1741 agccgccact agaggagcta ctgtagtaat tggaacaagc aaattctatg gtggttggca 1801 caacatgtta aaaactgttt atagtgatgt agaaaaccct caccttatgg gttgggatta 1861 tcctaaatgt gatagagcca tgcctaacat gcttagaatt atggcctcac ttgttcttgc 1921 tcgcaaacat acaacgtgtt gtagcttgtc acaccgtttc tatagattag ctaatgagtg 1981 tgctcaagta ttgagtgaaa tggtcatgtg tggcggttca ctatatgtta aaccaggtgg 2041 aacctcatca ggagatgcca caactgctta tgctaatagt gtttttaaca tttgtcaagc 2101 tgtcacggcc aatgttaatg cacttttatc tactgatggt aacaaaattg ccgataagta 2161 tgtccgcaat ttacaacaca gactttatga gtgtctctat agaaatagag atgttgacac 2221 agactttgtg aatgagtttt acgcatattt gcgtaaacat ttctcaatga tgatactctc 2281 tgacgatgct gttgtgtgtt tcaatagcac ttatgcatct caaggtctag tggctagcat 2341 aaagaacttt aagtcagttc tttattatca aaacaatgtt tttatgtctg aagcaaaatg 2401 ttggactgag actgacctta ctaaaggacc tcatgaattt tgctctcaac atacaatgct 2461 agttaaacag ggtgatgatt atgtgtacct tccttaccca gatccatcaa gaatcctagg 2521 ggccggctgt tttgtagatg atatcgtaaa aacagatggt acacttatga ttgaacggtt 2581 cgtgtcttta gctatagatg cttacccact tactaaacat cctaatcagg agtatgctga 2641 tgtctttcat ttgtacttac aatacataag aaagctacat gatgagttaa caggacacat 2701 gttagacatg tattctgtta tgcttactaa tgataacact tcaaggtatt gggaacctga 2761 gttttatgag gctatgtaca caccgcatac agtcttagaa aacctttact ttcagtcgca 2821 ccatcaccat caccattaag ctagcttgac tgactgagat acagcgtacc ttcagctcac 2881 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa 2941 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa 3001 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg 3061 ggaggttttt taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt 3121 atcgtagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg 3181 ggccggattg ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg 3241 tcttatactc ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact 3301 tccatacgtg tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg 3361 atctctcgat ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc 3421 agaagtagtt catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga 3481 aacttaatta accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta 3541 cgctcagaat tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat 3601 tacgagtttc atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa 3661 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg 3721 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc 3781 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc 3841 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc 3901 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg 3961 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc 4021 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga 4081 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc 4141 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac 4201 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg 4261 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc 4321 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa 4381 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta 4441 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt 4501 tgcatttaaa tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc 4561 gatccatctt gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg 4621 ccttggctat tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag 4681 atcgggatca cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg 4741 aggaatatcg aaatcggggc gcgcctggtg taccgagaac gatcctctca gtgcgagtct 4801 cgacgatcca tatcgttgct tggcagtcag ccagtcggaa tccagcttgg gacccaggaa 4861 gtccaatcgt cagatattgt actcaagcct ggtcacggca gcgtaccgat ctgtttaaac 4921 ctagatattg atagtctgat cggtcaacgt ataatcgagt cctagctttt gcaaacatct 4981 atcaagagac aggatcagca ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc 5041 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga 5101 aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg ttacatcgaa ctggatctca 5161 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ctttccaatg atgagcactt 5221 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg 5281 gtcgccgcat acactattct cagaatgact tggttgagta ttcaccagtc acagaaaagc 5341 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata 5401 acactgcggc caacttactt ctgacaacga ttggaggacc gaaggagcta accgcttttt 5461 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag 5521 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca accttgcgta 5581 aactattaac tggcgaacta cttactctag cttcccggca acagttgata gactggatgg 5641 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg 5701 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag 5761 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg 5821 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taaccgattc 5881 taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca 5941 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc 6001 cttaccagaa atttatcctt aagatcccga atcgtttaaa ctcgactctg gctctatcga 6061 atctccgtcg tttcgagctt acgcgaacag ccgtggcgct catttgctcg tcgggcatcg 6121 aatctcgtca gctatcgtca gcttaccttt ttggcagcga tcgcggctcc cgacatcttg 6181 gaccattagc tccacaggta tcttcttccc tctagtggtc ataacagcag cttcagctac 6241 ctctcaattc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctatcaa 6301 tcgttgcgtt acacacacaa aaaaccaaca cacatccatc ttcgatggat agcgatttta 6361 ttatctaact gctgatcgag tgtagccaga tctagtaatc aattacgggg tcattagttc 6421 atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac 6481 cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa 6541 tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag 6601 tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc 6661 ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct 6721 acgtattagt catcgctatt accatgctga tgcggttttg gcagtacatc aatgggcgtg 6781 gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt 6841 tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga 6901 cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga 6961 accgtcagat cagatctttg tcgatcctac catccactcg acacacccgc cagcggccgc //