LOCUS Exported 8378 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 helicase. ACCESSION . VERSION . KEYWORDS pGBW-m4133183 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8378) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 8378) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8378 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 1811..1831 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1832..1849 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 1918..2039 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1929..1948) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1983..2002 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2154..2200 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2230..2366 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2411..2482 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2708..3296) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2788..2807) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 3796..5005 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 4842..4861 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 5009..6037 /codon_start=1 /gene="aph(4)-Ia" /product="aminoglycoside phosphotransferase from E. coli" /label=HygR /note="confers resistance to hygromycin" /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA KE" polyA_signal 6064..6112 /note="synthetic polyadenylation signal" CDS 6378..7238 /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(6596..6615) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 7251..7387 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 7532..7603 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 7608..7654 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 7672..7720 /note="synthetic polyadenylation signal" enhancer 7801..8104 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 8106..8308 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 8259..8279 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 8377..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggcg gtgggggctt gcgtactatg taacagccag acgtcattgc gctgcggtgc 61 atgtataaga aggccgtttc tgtgttgcaa atgctgctac gatcatgtta tttcgacttc 121 tcacaagctt gtgctcagtg tcaatcctta tgtgtgtaat gcccccggat gcgacgtcac 181 agatgttacc caactgtacc tgggcggcat gtcctattat tgtaagagcc acaaaccacc 241 aatctccttc cccctgtgcg ccaacggaca ggtgtttggg ctctacaaaa acacatgtgt 301 gggatctgac aatgtcaccg acttcaacgc tatcgccaca tgcgattgga ctaatgcagg 361 cgactacatt ttagccaaca cctgtaccga acggctgaag ttatttgcgg ctgagactct 421 aaaggcaact gaggaaacct tcaaactttc atatggtatc gccacggtac gtgaggtgtt 481 gtctgatcga gaactgcatc tcagttggga ggtcgggaag cctagacctc cccttaatcg 541 aaattacgtg ttcacaggtt atagggttac caagaacagc aaagtccaga tcggcgaata 601 tacttttgag aagggagatt acggtgacgc cgtagtgtat cgtggcacaa caacatacaa 661 actgaatgtt ggggattact tcgtgctgac gtcccacacc gttatgccgc tctcagctcc 721 aactttggtt ccccaggaac attacgtgcg gattactgga ctctatccaa cgctaaacat 781 aagcgacgag ttttccagca acgtggctaa ttaccaaaag gtcggcatgc agaagtactc 841 cacccttcag ggtcctcctg ggactggcaa aagtcacttc gccatcggac tggcacttta 901 ctatcccagt gcgcgcattg tatataccgc ctgctcacat gcggcagtgg acgccctgtg 961 tgagaaggct ttaaagtatc tgccaataga caaatgctct cgcattatcc ctgccagggc 1021 cagagtggag tgctttgata agtttaaagt gaactcgacc ctcgaacagt atgtcttctg 1081 cacagttaac gcattgcccg aaacgaccgc cgatatagtt gtgttcgacg aaatctcaat 1141 ggctaccaat tatgatctct ccgtcgtcaa tgcacggctg cgggccaaac actacgtcta 1201 catcggggac cccgcgcagt taccagcacc gcgaacccta ctgactaagg gcacacttga 1261 gccagagtac ttcaacagcg tatgtagact gatgaaaacc attggccctg atatgttcct 1321 gggaacatgt cggaggtgcc ccgctgaaat agtggacacc gtttcagcac ttgtgtacga 1381 caataagctc aaagcacaca aggataaatc cgcacaatgt tttaagatgt tttacaaagg 1441 ggttattacc catgatgtgt ctagcgctat caatcgccct cagattggcg tggtgagaga 1501 gtttttgacg cgaaatcccg cctggcgaaa ggccgtattc atcagtcctt acaactcgca 1561 aaatgccgtc gctagtaaga ttttaggact ccctactcag acagttgata gctctcaagg 1621 ttctgaatac gattatgtga tttttactca gactaccgaa acggcacatt cttgtaacgt 1681 caaccgtttt aacgtggcca ttacaagggc caaggttggc atcttgtgta tcatgtcaga 1741 ccgggacctg tatgataaac tgcaattcac atctttggag ataccgcgcc gcaatgtcgc 1801 tacccttcag gaaaaccttt actttcagtc gcaccatcac catcaccatt aaggatccgc 1861 aggcctctgc tagcttgact gactgagata cagcgtacct tcagctcaca gacatgataa 1921 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt 1981 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta 2041 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt 2101 aaagcaagta aaacctctac aaatgtggta ttggcccatc tctatcggta tcgtagcata 2161 accccttggg gcctctaaac gggtcttgag gggttttttg tgcccctcgg gccggattgc 2221 tatctaccgg cattggcgca gaaaaaaatg cctgatgcga cgctgcgcgt cttatactcc 2281 cacatatgcc agattcagca acggatacgg cttccccaac ttgcccactt ccatacgtgt 2341 cctccttacc agaaatttat ccttaaggtc gtcagctatc ctgcaggcga tctctcgatt 2401 tcgatcaaga cattccttta atggtctttt ctggacacca ctaggggtca gaagtagttc 2461 atcaaacttt cttccctccc taatctcatt ggttaccttg ggctatcgaa acttaattaa 2521 ccagtcaagt cagctacttg gcgagatcga cttgtctggg tttcgactac gctcagaatt 2581 gcgtcagtca agttcgatct ggtccttgct attgcacccg ttctccgatt acgagtttca 2641 tttaaatcat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc 2701 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc 2761 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc 2821 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt 2881 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg 2941 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat 3001 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag 3061 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt 3121 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc 3181 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta 3241 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag 3301 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga 3361 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa 3421 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa 3481 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcatttaaat 3541 ttccgaactc tccaaggccc tcgtcggaaa atcttcaaac ctttcgtccg atccatcttg 3601 caggctacct ctcgaacgaa ctatcgcaag tctcttggcc ggccttgcgc cttggctatt 3661 gcttggcagc gcctatcgcc aggtattact ccaatcccga atatccgaga tcgggatcac 3721 ccgagagaag ttcaacctac atcctcaatc ccgatctatc cgagatccga ggaatatcga 3781 aatcggggcg cgcctggcct ccgcgccggg ttttggcgcc tcccgcgggc gcccccctcg 3841 tcacggcgag cgctgccacg tcagacgaag ggcgcaggag cgtcctgatc cttccgcccg 3901 gacgctcagg acagcggccc gctgctcata agactcggcc ttagaacccc agtatcagca 3961 gaaggacatt ttaggacggg acttgggtga ctctagggca ctggttttct ttccagagag 4021 cggaacaggc gaggaaaagt agtcccttct cggcgattct gcggagggat ctccgtgggg 4081 cggtgaacgc cgatgattat ataaggacgc gccgggtgtg gcacagctag ttccgtcgca 4141 gccgggattt gggtcgcggt tcttgtttgt ggatcgctgt gatcgtcact tggtgagtag 4201 cgggctgctg ggctggccgg ggctttcgtg gccgccgggc cgctcggtgg gacggaagcg 4261 tgtggagaga ccgccaaggg ctgtagtctg ggtccgcgag caaggttgcc ctgaactggg 4321 ggttgggggg agcgcagcaa aatggcggct gttcccgagt cttgaatgga agacgcttgt 4381 gaggcgggct gtgaggtcgt tgaaacaagg tggggggcat ggtgggcggc aagaacccaa 4441 ggtcttgagc ccttcgctaa tgcgggaaag ctcttattcg ggtgagatgg gctgggcacc 4501 atctggggac cctgacgtga agtttgtcac tgactggaga actcggtttg tcgtctgttg 4561 cgggggcggc agttatggcg gtgccgttgg gcagtgcacc cgtacctttg ggagcgcgcg 4621 ccctcgtcgt gtcgtgacgt cacccgttct gttggcttat aatgcagggt ggggccacct 4681 gccggtaggt gtgcggtagg cttttctccg tcgcaggacg cagggttcgg gcctagggta 4741 ggctctcctg aatcgacagg cgccggacct ctggtgaggg gagggataag tgaggcgtca 4801 gtttctttgg tcggttttat gtacctatct tcttaagtag ctgaagctcc ggttttgaac 4861 tatgcgctcg gggttggcga gtgtgttttg tgaagttttt taggcacctt ttgaaatgta 4921 atcatttggg tcaatatgta attttcagtg ttagacttgt aaattgtccg ctaaattctg 4981 gccgtttttg gcttttttgt tagacaacat gggtaaaaag cctgaactca ccgcgacgtc 5041 tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc agctctcgga 5101 gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg tcctgcgggt 5161 aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact ttgcatccgc 5221 cgcgctcccg attccggaag tgcttgacat tggggagttc agcgagagcc tgacctattg 5281 catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg aactgcccgc 5341 tgttctgcag ccggtcgcgg aggcaatgga tgccatcgct gccgccgatc ttagccagac 5401 gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat ggcgtgattt 5461 catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg acgacaccgt 5521 cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg actgccccga 5581 agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg acaatggccg 5641 cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat acgaggtcgc 5701 caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc gctacttcga 5761 gcggaggcat ccggagcttg caggatcgcc ccggctccgg gcgtatatgc tccgcattgg 5821 tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag cttgggcgca 5881 gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta cacaaatcgc 5941 ccgcagaagc gccgccgtct ggaccgatgg ctgtgtagaa gtactcgccg atagtggaaa 6001 ccgacgcccc agcactcgtc cgagggcaaa ggaataagct agtatgtaag cctagtctta 6061 gataataaaa tcgctatcca tcgaagatgg atgtgtgttg gttttttgtg tgtgtaacgc 6121 taggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg 6181 ttgcttggca gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat 6241 attgtactca agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt 6301 ctgatcggtc aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat 6361 cagcaggagg ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg 6421 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg 6481 aagatcagtt gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc 6541 ttgagagttt tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat 6601 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact 6661 attctcagaa tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca 6721 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact 6781 tacttctgac aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg 6841 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg 6901 agcgtgacac cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg 6961 aactacttac tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg 7021 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag 7081 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc 7141 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga 7201 tcgctgagat aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc 7261 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc 7321 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta 7381 tccttaagat cgtttaaact cgactctggc tctatcgaat ctccgtcgtt tcgagcttac 7441 gcgaacagcc gtggcgctca tttgctcgtc gggcatcgaa tctcgtcagc tatcgtcagc 7501 ttaccttttt ggcagcgatc gcggctcccg acatcttgga ccattagctc cacaggtatc 7561 ttcttccctc tagtggtcat aacagcagct tcagctacct ctcaattcaa aaaacccctc 7621 aagacccgtt tagaggcccc aaggggttat gctatcaatc gttgcgttac acacacaaaa 7681 aaccaacaca catccatctt cgatggatag cgattttatt atctaactgc tgatcgagtg 7741 tagccagatc tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg 7801 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt 7861 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca 7921 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc 7981 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta 8041 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac 8101 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg 8161 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg 8221 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt 8281 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc 8341 gatcctacca tccactcgac acacccgcca gcggccgc //