LOCUS Exported 7086 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 RNA-dependent RNA polymerase. ACCESSION . VERSION . KEYWORDS pGBW-m4134456 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 7086) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 7086) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..7086 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 2804..2824 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 2825..2902 /codon_start=1 /product="calmodulin-binding peptide" /label=CBP /note="derived from skeletal muscle myosin light chain kinase; binds calmodulin with nanomolar affinity in the presence of calcium" /translation="KRRWKKNFIAVSAANRFKKISSSGAL" polyA_signal 2955..3076 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2966..2985) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3020..3039 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 3191..3237 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 3267..3403 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3448..3519 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(3745..4333) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3825..3844) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 5079..5939 /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(5297..5316) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5952..6088 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6240..6311 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6316..6362 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6380..6428 /note="synthetic polyadenylation signal" enhancer 6509..6812 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 6814..7016 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6967..6987 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgtct gccgacgcac agtcgttcct gaatagagtt tgtggtgtat ccgcggcccg 61 gctaactcca tgcggaaccg ggacgagcac agatgtggtg taccgagctt ttgatatcta 121 taacgacaaa gtcgctggct tcgccaagtt tcttaagacc aactgttgca ggttccaaga 181 gaaagacgaa gatgataatc tcatagactc atactttgtg gttaagcgtc acacttttag 241 taattatcag catgaggaaa caatttataa cttgctgaag gactgccctg ccgtagcaaa 301 acacgacttc tttaaattcc gcatcgatgg cgatatggtc ccgcatatta gcagacagcg 361 gttaaccaag tacactatgg cagacctggt gtacgccctg cgacacttcg acgagggaaa 421 ctgtgacacg ttaaaagaaa tcctcgtgac ttataattgt tgcgatgacg attatttcaa 481 caagaaagat tggtacgatt ttgtcgagaa tcccgatata ctcagagtgt acgctaacct 541 gggggaaagg gttcgccagg cactattgaa gacagtccaa ttttgcgacg cgatgcggaa 601 tgccggcatt gtgggtgtcc tgacacttga taaccaggac ctgaatggca actggtatga 661 ctttggagat ttcattcaga ccacgcccgg gtccggagta cctgtcgttg actcttacta 721 ctcacttctg atgccaatcc taaccttaac ccgagctctc acagctgaga gtcacgtgga 781 caccgacctc acaaaaccgt atatcaagtg ggatttgctg aagtacgatt tcacggaaga 841 gaggttaaag ctctttgatc gttatttcaa atactgggac cagacttatc atcccaactg 901 tgtgaattgt ctggacgaca gatgcatact tcattgtgcc aatttcaatg tactgtttag 961 cacagtgttc ccaccaactt cgtttggtcc tctagtgcgc aaaatctttg tggatggggt 1021 gcccttcgtt gtttccacag gctatcactt tcgtgagctc ggcgtagttc acaaccaaga 1081 tgtcaacctg cattcttcca ggttaagttt taaggaactt ctggtgtatg ccgcagaccc 1141 tgcgatgcac gctgcaagcg gtaatttgtt gctggataag cggaccacgt gcttcagcgt 1201 ggcggcttta acaaataacg ttgccttcca aaccgtcaag cctggcaact tcaataaaga 1261 tttctacgat ttcgccgtca gtaaagggtt ttttaaggag ggatcaagcg tagaattgaa 1321 acattttttc tttgctcagg acggtaatgc cgctatctcg gactacgact actaccggta 1381 taacctacca actatgtgcg acattcggca gctgttgttc gtggtggagg tcgttgataa 1441 atactttgat tgctacgatg gcggatgcat taacgccaat caagtcatcg tgaacaatct 1501 ggacaagagc gccggcttcc cctttaacaa atggggaaag gcaaggctat attatgactc 1561 tatgtcctat gaggatcagg acgctctctt cgcgtatacc aagcgcaacg ttatccccac 1621 aattacccag atgaacctca aatatgcaat cagcgcaaag aacagagcga ggacagttgc 1681 tggggtgagt atatgttcga caatgactaa caggcagttt caccagaagc tgttgaaatc 1741 tattgccgct acacgagggg ctaccgtcgt gattggcacc tccaaattct acggagggtg 1801 gcataatatg ttaaaaactg tgtactccga cgtggaaaac cctcacctga tgggttggga 1861 ttatcctaag tgcgaccggg cgatgccaaa catgctgaga attatggcgt cattagtcct 1921 ggcgcgaaag cacaccacgt gctgtagtct gtcacatcgc ttttacagat tggctaatga 1981 atgcgcccaa gtgcttagcg agatggtgat gtgtgggggt tcactttacg tgaagcccgg 2041 aggaacgtcc tcaggcgacg ccacaactgc ctacgcaaat tcagtattca acatatgtca 2101 ggccgttaca gcaaacgtga atgctctgct gtctactgat gggaacaaga tcgccgataa 2161 gtacgtccgg aacctgcaac acaggctgta cgagtgtctg tatcgtaatc gagatgtaga 2221 caccgacttc gtgaacgaat tttacgcata cttacggaaa catttctcca tgatgatcct 2281 ctccgacgat gcagttgtgt gctttaattc aacttacgcc tctcagggct tggtggccag 2341 tatcaaaaat tttaagtcgg ttctttatta tcagaataac gtgtttatga gcgaggctaa 2401 gtgttggacc gagactgatc tgaccaaagg cccgcacgag ttctgttccc agcacaccat 2461 gctggtcaag cagggtgatg actatgtcta tttaccatac ccagatccga gtaggatctt 2521 gggagccggt tgttttgtgg atgacattgt gaagactgac ggtacactca tgattgagcg 2581 cttcgtcagt ttggcaatcg acgcttaccc ccttaccaaa catccaaatc aggaatatgc 2641 ggatgtgttt cacctgtacc ttcaatacat aaggaaactt catgatgaac taactggaca 2701 tatgctcgac atgtattctg taatgcttac aaatgacaat acctcacggt actgggaacc 2761 ggagttttat gaagccatgt acaccccaca tactgtcctg caagaaaacc tttactttca 2821 gtcgaagagg cgatggaaaa agaattttat cgcggtgagt gcagccaaca gattcaaaaa 2881 gatatcctcg tctggtgccc tctaagctag cttgactgac tgagatacag cgtaccttca 2941 gctcacagac atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg 3001 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag 3061 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga 3121 ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtattg gcccatctct 3181 atcggtatcg tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgtgc 3241 ccctcgggcc ggattgctat ctaccggcat tggcgcagaa aaaaatgcct gatgcgacgc 3301 tgcgcgtctt atactcccac atatgccaga ttcagcaacg gatacggctt ccccaacttg 3361 cccacttcca tacgtgtcct ccttaccaga aatttatcct taaggtcgtc agctatcctg 3421 caggcgatct ctcgatttcg atcaagacat tcctttaatg gtcttttctg gacaccacta 3481 ggggtcagaa gtagttcatc aaactttctt ccctccctaa tctcattggt taccttgggc 3541 tatcgaaact taattaacca gtcaagtcag ctacttggcg agatcgactt gtctgggttt 3601 cgactacgct cagaattgcg tcagtcaagt tcgatctggt ccttgctatt gcacccgttc 3661 tccgattacg agtttcattt aaatcatgtg agcaaaaggc cagcaaaagg ccaggaaccg 3721 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa 3781 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt 3841 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct 3901 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct 3961 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc 4021 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt 4081 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc 4141 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat 4201 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa 4261 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa 4321 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga 4381 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct 4441 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga 4501 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc 4561 catagttgca tttaaatttc cgaactctcc aaggccctcg tcggaaaatc ttcaaacctt 4621 tcgtccgatc catcttgcag gctacctctc gaacgaacta tcgcaagtct cttggccggc 4681 cttgcgcctt ggctattgct tggcagcgcc tatcgccagg tattactcca atcccgaata 4741 tccgagatcg ggatcacccg agagaagttc aacctacatc ctcaatcccg atctatccga 4801 gatccgagga atatcgaaat cggggcgcgc ctggtgtacc gagaacgatc ctctcagtgc 4861 gagtctcgac gatccatatc gttgcttggc agtcagccag tcggaatcca gcttgggacc 4921 caggaagtcc aatcgtcaga tattgtactc aagcctggtc acggcagcgt accgatctgt 4981 ttaaacctag atattgatag tctgatcggt caacgtataa tcgagtccta gcttttgcaa 5041 acatctatca agagacagga tcagcaggag gctttcgcat gagtattcaa catttccgtg 5101 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc 5161 tggtgaaagt aaaagatgct gaagatcagt tgggtgcgcg agtgggttac atcgaactgg 5221 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgcttt ccaatgatga 5281 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc 5341 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtattca ccagtcacag 5401 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga 5461 gtgataacac tgcggccaac ttacttctga caacgattgg aggaccgaag gagctaaccg 5521 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga 5581 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacct 5641 tgcgtaaact attaactggc gaactactta ctctagcttc ccggcaacag ttgatagact 5701 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt 5761 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg 5821 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta 5881 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac 5941 cgattctagg tgcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact 6001 cccacatatg ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt 6061 gtcctcctta ccagaaattt atccttaaga tcccgaatcg tttaaactcg actctggctc 6121 tatcgaatct ccgtcgtttc gagcttacgc gaacagccgt ggcgctcatt tgctcgtcgg 6181 gcatcgaatc tcgtcagcta tcgtcagctt acctttttgg cagcgatcgc ggctcccgac 6241 atcttggacc attagctcca caggtatctt cttccctcta gtggtcataa cagcagcttc 6301 agctacctct caattcaaaa aacccctcaa gacccgttta gaggccccaa ggggttatgc 6361 tatcaatcgt tgcgttacac acacaaaaaa ccaacacaca tccatcttcg atggatagcg 6421 attttattat ctaactgctg atcgagtgta gccagatcta gtaatcaatt acggggtcat 6481 tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg 6541 gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa 6601 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact 6661 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta 6721 aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt 6781 acatctacgt attagtcatc gctattacca tgctgatgcg gttttggcag tacatcaatg 6841 ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg 6901 ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc 6961 cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctggtt 7021 tagtgaaccg tcagatcaga tctttgtcga tcctaccatc cactcgacac acccgccagc 7081 ggccgc //