LOCUS Exported 6666 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp11. ACCESSION . VERSION . KEYWORDS pGBW-m4252928 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6666) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6666) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6666 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 233..821 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 722..741 /label=pBR322ori-F /note="pBR322 origin, forward primer" misc_feature 1047..1118 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" terminator 1163..1299 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" terminator 1329..1375 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 1490..1611 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1527..1546) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 1581..1600 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" CDS complement(1680..1745) /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" CDS complement(1749..1769) /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" regulatory 1802..1811 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" promoter complement(1880..2082) /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind complement(1909..1929) /label=CMV-F /note="Human CMV immediate early promoter, forward primer" enhancer 2084..2387 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" polyA_signal 2468..2516 /note="synthetic polyadenylation signal" terminator 2534..2580 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" misc_feature 2585..2656 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 2808..2944 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" CDS complement(2957..3817) /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 3580..3599 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" polyA_signal 4083..4131 /note="synthetic polyadenylation signal" CDS complement(4158..5186) /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" promoter complement(5190..6399) /label=UbC promoter /note="human ubiquitin C promoter" primer_bind complement(5334..5353) /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" ORIGIN 1 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt 61 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat 121 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg 181 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc 241 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg 301 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag 361 cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact 421 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg 481 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc 541 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg 601 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg 661 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag 721 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc 781 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct 841 ttttacggtt cctggccttt tgctggcctt ttgctcacat gatttaaatg aaactcgtaa 901 tcggagaacg ggtgcaatag caaggaccag atcgaacttg actgacgcaa ttctgagcgt 961 agtcgaaacc cagacaagtc gatctcgcca agtagctgac ttgactggtt aattaagttt 1021 cgatagccca aggtaaccaa tgagattagg gagggaagaa agtttgatga actacttctg 1081 acccctagtg gtgtccagaa aagaccatta aaggaatgtc ttgatcgaaa tcgagagatc 1141 gcctgcagga tagctgacga ccttaaggat aaatttctgg taaggaggac acgtatggaa 1201 gtgggcaagt tggggaagcc gtatccgttg ctgaatctgg catatgtggg agtataagac 1261 gcgcagcgtc gcatcaggca tttttttctg cgccaatgcc ggtagatagc aatccggccc 1321 gaggggcaca aaaaacccct caagacccgt ttagaggccc caaggggtta tgctacgata 1381 ccgatagaga tgggccaata ccacatttgt agaggtttta cttgctttaa aaaacctccc 1441 acacctcccc ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat 1501 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt 1561 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg 1621 tgagctgaag gtacgctgta tctcagtcag tcaagctagc agaggcctgc ggatccttat 1681 ttgtcatcgt cgtccttgta atcaatgtca tgatctttgt agtctccatc gtggtcctta 1741 taatccatcg actgaaagta aaggttttcc gcaaacccgt ttaaaaacga ttgtgcatca 1801 gccatggtgg cggccgctgg cgggtgtgtc gagtggatgg taggatcgac aaagatctga 1861 tctgacggtt cactaaacca gctctgctta tatagacctc ccaccgtaca cgcctaccgc 1921 ccatttgcgt caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt 1981 gccaaaacaa actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa 2041 accgctatcc acgcccattg atgtactgcc aaaaccgcat cagcatggta atagcgatga 2101 ctaatacgta gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat 2161 gccaggcggg ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca 2221 cttgatgtac tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga 2281 aagtccctat tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt 2341 cgttgggcgg tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat 2401 atgggctatg aactaatgac cccgtaattg attactagat ctggctacac tcgatcagca 2461 gttagataat aaaatcgcta tccatcgaag atggatgtgt gttggttttt tgtgtgtgta 2521 acgcaacgat tgatagcata accccttggg gcctctaaac gggtcttgag gggttttttg 2581 aattgagagg tagctgaagc tgctgttatg accactagag ggaagaagat acctgtggag 2641 ctaatggtcc aagatgtcgg gagccgcgat cgctgccaaa aaggtaagct gacgatagct 2701 gacgagattc gatgcccgac gagcaaatga gcgccacggc tgttcgcgta agctcgaaac 2761 gacggagatt cgatagagcc agagtcgagt ttaaacgatt cgggatctta aggataaatt 2821 tctggtaagg aggacacgta tggaagtggg caagttgggg aagccgtatc cgttgctgaa 2881 tctggcatat gtgggagtat aagacgcgca gcgtcgcatc aggcattttt ttctgcgcca 2941 atgcacctag aatcggttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc 3001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg 3061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag 3121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt 3181 tatccgcctc catccagtct atcaactgtt gccgggaagc tagagtaagt agttcgccag 3241 ttaatagttt acgcaaggtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt 3301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca 3361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccaat cgttgtcaga agtaagttgg 3421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat 3481 ccgtaagatg cttttctgtg actggtgaat actcaaccaa gtcattctga gaatagtgta 3541 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca 3601 gaactttaaa agtgctcatc attggaaagc gttcttcggg gcgaaaactc tcaaggatct 3661 taccgctgtt gagatccagt tcgatgtaac ccactcgcgc acccaactga tcttcagcat 3721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa 3781 agggaataag ggcgacacgg aaatgttgaa tactcatgcg aaagcctcct gctgatcctg 3841 tctcttgata gatgtttgca aaagctagga ctcgattata cgttgaccga tcagactatc 3901 aatatctagg tttaaacaga tcggtacgct gccgtgacca ggcttgagta caatatctga 3961 cgattggact tcctgggtcc caagctggat tccgactggc tgactgccaa gcaacgatat 4021 ggatcgtcga gactcgcact gagaggatcg ttctcggtac accaggcgcg cctagcgtta 4081 cacacacaaa aaaccaacac acatccatct tcgatggata gcgattttat tatctaagac 4141 taggcttaca tactagctta ttcctttgcc ctcggacgag tgctggggcg tcggtttcca 4201 ctatcggcga gtacttctac acagccatcg gtccagacgg cggcgcttct gcgggcgatt 4261 tgtgtacgcc cgacagtccc ggctccggat cggacgattg cgtcgcatcg accctgcgcc 4321 caagctgcat catcgaaatt gccgtcaacc aagctctgat agagttggtc aagaccaatg 4381 cggagcatat acgcccggag ccggggcgat cctgcaagct ccggatgcct ccgctcgaag 4441 tagcgcgtct gctgctccat acaagccaac cacggcctcc agaagaagat gttggcgacc 4501 tcgtattggg aatccccgaa catcgcctcg ctccagtcaa tgaccgctgt tatgcggcca 4561 ttgtccgtca ggacattgtt ggagccgaaa tccgcgtgca cgaggtgccg gacttcgggg 4621 cagtcctcgg cccaaagcat cagctcatcg agagcctgcg cgacggacgc actgacggtg 4681 tcgtccatca cagtttgcca gtgatacaca tggggatcag caatcgcgca tatgaaatca 4741 cgccatgtag tgtattgacc gattccttgc ggtccgaatg ggccgaaccc gctcgtctgg 4801 ctaagatcgg cggcagcgat ggcatccatt gcctccgcga ccggctgcag aacagcgggc 4861 agttcggttt caggcaggtc ttgcaacgtg acaccctgtg cacggcggga gatgcaatag 4921 gtcaggctct cgctgaactc cccaatgtca agcacttccg gaatcgggag cgcggcggat 4981 gcaaagtgcc gataaacata acgatctttg tagaaaccat cggcgcagct atttacccgc 5041 aggacatatc cacgccctcc tacatcgaag ctgaaagcac gagattcttc gccctccgag 5101 agctgcatca ggtcggagac gctgtcgaac ttttcgatca gaaacttctc gacagacgtc 5161 gcggtgagtt caggcttttt acccatgttg tctaacaaaa aagccaaaaa cggccagaat 5221 ttagcggaca atttacaagt ctaacactga aaattacata ttgacccaaa tgattacatt 5281 tcaaaaggtg cctaaaaaac ttcacaaaac acactcgcca accccgagcg catagttcaa 5341 aaccggagct tcagctactt aagaagatag gtacataaaa ccgaccaaag aaactgacgc 5401 ctcacttatc cctcccctca ccagaggtcc ggcgcctgtc gattcaggag agcctaccct 5461 aggcccgaac cctgcgtcct gcgacggaga aaagcctacc gcacacctac cggctggtgg 5521 ccccaccctg cattataagc caacagaacg ggtgacgtca cgacacgacg agggcgcgcg 5581 ctcccaaagg tacgggtgca ctgcccaacg gcaccgccat aactgccgcc cccgcaacag 5641 acgacaaacc gagttctcca gtcagtgaca aacttcacgt cagggtcccc agatggtgcc 5701 cagcccatct cacccgaata agagctttcc cgcattagcg aagggctcaa gaccttgggt 5761 tcttgccgcc caccatgccc cccaccttgt ttcaacgacc tcacagcccg cctcacaagc 5821 gtcttccatt caagactcgg gaacagccgc cattttgctg cgctcccccc aacccccagt 5881 tcagggcaac cttgctcgcg gacccagact acagcccttg gcggtctctc cacacgcttc 5941 cgtcccaccg agcggcccgg cggccacgaa agccccggcc agcccagcag cccgctactc 6001 accaagtgac gatcacagcg atccacaaac aagaaccgcg acccaaatcc cggctgcgac 6061 ggaactagct gtgccacacc cggcgcgtcc ttatataatc atcggcgttc accgccccac 6121 ggagatccct ccgcagaatc gccgagaagg gactactttt cctcgcctgt tccgctctct 6181 ggaaagaaaa ccagtgccct agagtcaccc aagtcccgtc ctaaaatgtc cttctgctga 6241 tactggggtt ctaaggccga gtcttatgag cagcgggccg ctgtcctgag cgtccgggcg 6301 gaaggatcag gacgctcctg cgcccttcgt ctgacgtggc agcgctcgcc gtgacgaggg 6361 gggcgcccgc gggaggcgcc aaaacccggc gcggaggcca ggcgcgcccc gatttcgata 6421 ttcctcggat ctcggataga tcgggattga ggatgtaggt tgaacttctc tcgggtgatc 6481 ccgatctcgg atattcggga ttggagtaat acctggcgat aggcgctgcc aagcaatagc 6541 caaggcgcaa ggccggccaa gagacttgcg atagttcgtt cgagaggtag cctgcaagat 6601 ggatcggacg aaaggtttga agattttccg acgagggcct tggagagttc ggaaatttaa 6661 atgcaa //