LOCUS Exported 5685 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 3C-like proteinase. ACCESSION . VERSION . KEYWORDS pGBW-m4133345 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5685) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5685) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5685 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 926..946 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 980..1057 /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" CDS 1085..1105 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQG" polyA_signal 1554..1675 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1565..1584) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1619..1638 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1790..1836 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1866..2002 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2047..2118 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2344..2932) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2424..2443) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 3678..4538 /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(3896..3915) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 4551..4687 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 4839..4910 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4915..4961 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 4979..5027 /note="synthetic polyadenylation signal" enhancer 5108..5411 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 5413..5615 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 5566..5586 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgagt gggttccgga aaatggcgtt tccctccggt aaggtcgaag gatgcatggt 61 tcaggtgacc tgtggcacaa ctacgttaaa cggcctatgg ctggacgatg tggtatactg 121 tcctcgacac gtcatctgca catcagagga catgctgaat ccaaactatg aggatttgct 181 cataagaaag agcaatcata actttctggt gcaagctgga aatgtacagc ttagggttat 241 tgggcactct atgcagaact gcgtgttaaa actcaaggtg gataccgcca atccgaaaac 301 acctaagtac aaattcgtgc gcatccagcc aggccaaacc ttctctgtcc tggcatgtta 361 taacggcagc ccctcaggtg tctaccagtg cgccatgcgc cctaatttta ctattaaggg 421 ttccttcttg aacgggtcgt gcggatcggt cggctttaat atcgactatg actgtgtgag 481 tttttgctac atgcatcaca tggaacttcc gactggggtt cacgctggaa cggatctgga 541 gggtaacttc tatggaccct tcgtggatag acagaccgcc caggcagccg gcactgacac 601 taccattacg gtgaatgttc tagcttggct ctacgcagct gtaataaacg gcgatcgttg 661 gttcttgaac aggtttacaa caaccctgaa cgactttaat ctagtcgcca tgaaatataa 721 ttatgaacca ctgacccaag atcatgtaga cattctcggg ccactgagcg cacagacagg 781 gatcgctgtg ctggacatgt gtgccagttt aaaggagctg cttcagaatg gtatgaacgg 841 ccgaacaatc ctagggtctg cgctcttaga ggatgaattt acccccttcg atgtagtgcg 901 gcaatgtagc ggcgttactt ttcaagaaaa cctttacttt cagtcgggac ggcgaatccc 961 ggggctgata aatccctgga aaaggagatg gaagaagaac tttattgcag tgtctgcggc 1021 taaccggttc aaaaagatca gttcaagcgg cgccctagat tacgacattc ctacgaccgc 1081 ctccgagaat ctctatttcc agggtgaatt tggcttggca caacatgacg aggccgtcga 1141 taacaagttt aataaagaac agcagaacgc tttctatgag atccttcact taccaaatct 1201 gaacgaagag caacgcaacg ctttcattca gagcctgaaa gacgatccct cgcagtctgc 1261 aaatctcctg gccgaagcca agaagctgaa tgacgcacag gctccaaaag ttgataacaa 1321 gtttaacaag gagcagcaga atgctttcta cgagatattg caccttccta atctgaatga 1381 ggaacaaaga aatgcattta tccagtccct aaaagacgat ccatcacaga gtgccaacct 1441 gcttgcggaa gccaaaaagc tcaatgatgc ccaggccccg aaagtagacg cgaaccatca 1501 ataagctagc ttgactgact gagatacagc gtaccttcag ctcacagaca tgataagata 1561 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga 1621 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa 1681 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag 1741 caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt agcataaccc 1801 cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg gattgctatc 1861 taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca 1921 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc 1981 cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc tcgatttcga 2041 tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag tagttcatca 2101 aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt aattaaccag 2161 tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc agaattgcgt 2221 cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga gtttcattta 2281 aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc 2341 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag 2401 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt 2461 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg 2521 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg 2581 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg 2641 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac 2701 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg 2761 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt 2821 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg 2881 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc 2941 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt 3001 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt 3061 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag 3121 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat ttaaatttcc 3181 gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc atcttgcagg 3241 ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg gctattgctt 3301 ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg gatcacccga 3361 gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa tatcgaaatc 3421 ggggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg 3481 ttgcttggca gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat 3541 attgtactca agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt 3601 ctgatcggtc aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat 3661 cagcaggagg ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg 3721 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg 3781 aagatcagtt gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc 3841 ttgagagttt tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat 3901 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact 3961 attctcagaa tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca 4021 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact 4081 tacttctgac aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg 4141 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg 4201 agcgtgacac cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg 4261 aactacttac tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg 4321 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag 4381 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc 4441 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga 4501 tcgctgagat aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc 4561 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc 4621 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta 4681 tccttaagat cccgaatcgt ttaaactcga ctctggctct atcgaatctc cgtcgtttcg 4741 agcttacgcg aacagccgtg gcgctcattt gctcgtcggg catcgaatct cgtcagctat 4801 cgtcagctta cctttttggc agcgatcgcg gctcccgaca tcttggacca ttagctccac 4861 aggtatcttc ttccctctag tggtcataac agcagcttca gctacctctc aattcaaaaa 4921 acccctcaag acccgtttag aggccccaag gggttatgct atcaatcgtt gcgttacaca 4981 cacaaaaaac caacacacat ccatcttcga tggatagcga ttttattatc taactgctga 5041 tcgagtgtag ccagatctag taatcaatta cggggtcatt agttcatagc ccatatatgg 5101 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc 5161 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt 5221 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc 5281 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg 5341 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg 5401 ctattaccat gctgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact 5461 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa 5521 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta 5581 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatcagat 5641 ctttgtcgat cctaccatcc actcgacaca cccgccagcg gccgc //