LOCUS Exported 8388 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 helicase. ACCESSION . VERSION . KEYWORDS pGBW-m4133829 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 8388) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 8388) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..8388 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 81..384 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 386..588 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 539..559 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 657..666 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2463..2483 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 2484..2510 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" polyA_signal 2579..2700 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2590..2609) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2644..2663 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2815..2861 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2891..3027 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3072..3143 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(3369..3957) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3449..3468) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 4457..5666 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 5503..5522 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 5670..6698 /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 6725..6773 /note="synthetic polyadenylation signal" CDS 7039..7899 /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(7257..7276) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 7912..8048 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 8200..8271 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 8276..8322 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 8340..8388 /note="synthetic polyadenylation signal" ORIGIN 1 atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc attagttcat 61 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg 121 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata 181 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta 241 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc 301 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac 361 gtattagtca tcgctattac catgctgatg cggttttggc agtacatcaa tgggcgtgga 421 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg 481 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg 541 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg tttagtgaac 601 cgtcagatca gatctttgtc gatcctacca tccactcgac acacccgcca gcggccgcca 661 ccatggttgg ggcttgtgtt ctttgcaatt cacagacttc attaagatgt ggtgcttgca 721 tacgtagacc attcttatgt tgtaaatgct gttacgacca tgtcatatca acatcacata 781 aattagtctt gtctgttaat ccgtatgttt gcaatgctcc aggttgtgat gtcacagatg 841 tgactcaact ttacttagga ggtatgagct attattgtaa atcacataaa ccacccatta 901 gttttccatt gtgtgctaat ggacaagttt ttggtttata taaaaataca tgtgttggta 961 gcgataatgt tactgacttt aatgcaattg caacatgtga ctggacaaat gctggtgatt 1021 acattttagc taacacctgt actgaaagac tcaagctttt tgcagcagaa acgctcaaag 1081 ctactgagga gacatttaaa ctgtcttatg gtattgctac tgtacgtgaa gtgctgtctg 1141 acagagaatt acatctttca tgggaagttg gtaaacctag accaccactt aaccgaaatt 1201 atgtctttac tggttatcgt gtaactaaaa acagtaaagt acaaatagga gagtacacct 1261 ttgaaaaagg tgactatggt gatgctgttg tttaccgagg tacaacaact tacaaattaa 1321 atgttggtga ttattttgtg ctgacatcac atacagtaat gccattaagt gcacctacac 1381 tagtgccaca agagcactat gttagaatta ctggcttata cccaacactc aatatctcag 1441 atgagttttc tagcaatgtt gcaaattatc aaaaggttgg tatgcaaaag tattctacac 1501 tccagggacc acctggtact ggtaagagtc attttgctat tggcctagct ctctactacc 1561 cttctgctcg catagtgtat acagcttgct ctcatgccgc tgttgatgca ctatgtgaga 1621 aggcattaaa atatttgcct atagataaat gtagtagaat tatacctgca cgtgctcgtg 1681 tagagtgttt tgataaattc aaagtgaatt caacattaga acagtatgtc ttttgtactg 1741 taaatgcatt gcctgagacg acagcagata tagttgtctt tgatgaaatt tcaatggcca 1801 caaattatga tttgagtgtt gtcaatgcca gattacgtgc taagcactat gtgtacattg 1861 gcgaccctgc tcaattacct gcaccacgca cattgctaac taagggcaca ctagaaccag 1921 aatatttcaa ttcagtgtgt agacttatga aaactatagg tccagacatg ttcctcggaa 1981 cttgtcggcg ttgtcctgct gaaattgttg acactgtgag tgctttggtt tatgataata 2041 agcttaaagc acataaagac aaatcagctc aatgctttaa aatgttttat aagggtgtta 2101 tcacgcatga tgtttcatct gcaattaaca ggccacaaat aggcgtggta agagaattcc 2161 ttacacgtaa ccctgcttgg agaaaagctg tctttatttc accttataat tcacagaatg 2221 ctgtagcctc aaagattttg ggactaccaa ctcaaactgt tgattcatca cagggctcag 2281 aatatgacta tgtcatattc actcaaacca ctgaaacagc tcactcttgt aatgtaaaca 2341 gatttaatgt tgctattacc agagcaaaag taggcatact ttgcataatg tctgatagag 2401 acctttatga caagttgcaa tttacaagtc ttgaaattcc acgtaggaat gtggcaactt 2461 tagaaaacct ttactttcag tcgtacccct atgacgtacc agattatgcg taaggatccg 2521 caggcctctg ctagcttgac tgactgagat acagcgtacc ttcagctcac agacatgata 2581 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt 2641 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt 2701 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt 2761 taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt atcgtagcat 2821 aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg ggccggattg 2881 ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc 2941 ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg 3001 tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg atctctcgat 3061 ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc agaagtagtt 3121 catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga aacttaatta 3181 accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta cgctcagaat 3241 tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat tacgagtttc 3301 atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg 3361 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt 3421 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc 3481 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct 3541 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc 3601 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta 3661 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca 3721 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag 3781 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag 3841 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt 3901 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa 3961 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg 4021 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga 4081 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta 4141 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcatttaaa 4201 tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt 4261 gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg ccttggctat 4321 tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag atcgggatca 4381 cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg aggaatatcg 4441 aaatcggggc gcgcctggcc tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc 4501 gtcacggcga gcgctgccac gtcagacgaa gggcgcagga gcgtcctgat ccttccgccc 4561 ggacgctcag gacagcggcc cgctgctcat aagactcggc cttagaaccc cagtatcagc 4621 agaaggacat tttaggacgg gacttgggtg actctagggc actggttttc tttccagaga 4681 gcggaacagg cgaggaaaag tagtcccttc tcggcgattc tgcggaggga tctccgtggg 4741 gcggtgaacg ccgatgatta tataaggacg cgccgggtgt ggcacagcta gttccgtcgc 4801 agccgggatt tgggtcgcgg ttcttgtttg tggatcgctg tgatcgtcac ttggtgagta 4861 gcgggctgct gggctggccg gggctttcgt ggccgccggg ccgctcggtg ggacggaagc 4921 gtgtggagag accgccaagg gctgtagtct gggtccgcga gcaaggttgc cctgaactgg 4981 gggttggggg gagcgcagca aaatggcggc tgttcccgag tcttgaatgg aagacgcttg 5041 tgaggcgggc tgtgaggtcg ttgaaacaag gtggggggca tggtgggcgg caagaaccca 5101 aggtcttgag cccttcgcta atgcgggaaa gctcttattc gggtgagatg ggctgggcac 5161 catctgggga ccctgacgtg aagtttgtca ctgactggag aactcggttt gtcgtctgtt 5221 gcgggggcgg cagttatggc ggtgccgttg ggcagtgcac ccgtaccttt gggagcgcgc 5281 gccctcgtcg tgtcgtgacg tcacccgttc tgttggctta taatgcaggg tggggccacc 5341 tgccggtagg tgtgcggtag gcttttctcc gtcgcaggac gcagggttcg ggcctagggt 5401 aggctctcct gaatcgacag gcgccggacc tctggtgagg ggagggataa gtgaggcgtc 5461 agtttctttg gtcggtttta tgtacctatc ttcttaagta gctgaagctc cggttttgaa 5521 ctatgcgctc ggggttggcg agtgtgtttt gtgaagtttt ttaggcacct tttgaaatgt 5581 aatcatttgg gtcaatatgt aattttcagt gttagacttg taaattgtcc gctaaattct 5641 ggccgttttt ggcttttttg ttagacaaca tgggtaaaaa gcctgaactc accgcgacgt 5701 ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg 5761 agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg 5821 taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatccg 5881 ccgcgctccc gattccggaa gtgcttgaca ttggggagtt cagcgagagc ctgacctatt 5941 gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg 6001 ctgttctgca gccggtcgcg gaggcaatgg atgccatcgc tgccgccgat cttagccaga 6061 cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt 6121 tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg 6181 tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg 6241 aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc 6301 gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg 6361 ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg 6421 agcggaggca tccggagctt gcaggatcgc cccggctccg ggcgtatatg ctccgcattg 6481 gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc 6541 agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg 6601 cccgcagaag cgccgccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa 6661 accgacgccc cagcactcgt ccgagggcaa aggaataagc tagtatgtaa gcctagtctt 6721 agataataaa atcgctatcc atcgaagatg gatgtgtgtt ggttttttgt gtgtgtaacg 6781 ctaggcgcgc ctggtgtacc gagaacgatc ctctcagtgc gagtctcgac gatccatatc 6841 gttgcttggc agtcagccag tcggaatcca gcttgggacc caggaagtcc aatcgtcaga 6901 tattgtactc aagcctggtc acggcagcgt accgatctgt ttaaacctag atattgatag 6961 tctgatcggt caacgtataa tcgagtccta gcttttgcaa acatctatca agagacagga 7021 tcagcaggag gctttcgcat gagtattcaa catttccgtg tcgcccttat tccctttttt 7081 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct 7141 gaagatcagt tgggtgcgcg agtgggttac atcgaactgg atctcaacag cggtaagatc 7201 cttgagagtt ttcgccccga agaacgcttt ccaatgatga gcacttttaa agttctgcta 7261 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac 7321 tattctcaga atgacttggt tgagtattca ccagtcacag aaaagcatct tacggatggc 7381 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac 7441 ttacttctga caacgattgg aggaccgaag gagctaaccg cttttttgca caacatgggg 7501 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac 7561 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacct tgcgtaaact attaactggc 7621 gaactactta ctctagcttc ccggcaacag ttgatagact ggatggaggc ggataaagtt 7681 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga 7741 gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc 7801 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag 7861 atcgctgaga taggtgcctc actgattaag cattggtaac cgattctagg tgcattggcg 7921 cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag 7981 caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt 8041 atccttaaga tcccgaatcg tttaaactcg actctggctc tatcgaatct ccgtcgtttc 8101 gagcttacgc gaacagccgt ggcgctcatt tgctcgtcgg gcatcgaatc tcgtcagcta 8161 tcgtcagctt acctttttgg cagcgatcgc ggctcccgac atcttggacc attagctcca 8221 caggtatctt cttccctcta gtggtcataa cagcagcttc agctacctct caattcaaaa 8281 aacccctcaa gacccgttta gaggccccaa ggggttatgc tatcaatcgt tgcgttacac 8341 acacaaaaaa ccaacacaca tccatcttcg atggatagcg attttatt //