LOCUS Exported 6987 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp11. ACCESSION . VERSION . KEYWORDS pGBW-m4252897 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6987) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6987) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6987 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 41..61 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 410..427 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 520..641 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(531..550) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 585..604 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 756..802 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 832..968 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1013..1084 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1310..1898) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1390..1409) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 2398..3607 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3444..3463 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 3611..4639 /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 4666..4714 /note="synthetic polyadenylation signal" CDS 4980..5840 /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(5198..5217) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5853..5989 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 6141..6212 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 6217..6263 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 6281..6329 /note="synthetic polyadenylation signal" enhancer 6410..6713 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 6715..6917 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6868..6888 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 6986..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggct gatgcacaat cgtttttaaa cgggtttgcg gaaaaccttt actttcagtc 61 gatgagtgac aaaatcattc atctaaccga tgactccttt gatacagacg tgttgaaggc 121 cgatggtgcg atactggtag atttctgggc tgaatggtgt ggcccgtgca agatgatcgc 181 acccattctg gacgagatcg ccgacgagta tcaaggaaaa ttaactgtcg ctaagcttaa 241 tatagatcag aacccaggga cggcacctaa atacggcatc aggggaatac ctaccctcct 301 gctctttaag aatggggaag ttgccgcgac aaaagtggga gccctgtcta agggccagct 361 aaaagagttc ttggacgcta acctggcagg ttcaggctcg gggcacatgc accatcacca 421 tcaccatagc agcgggctcg tgccacgcgg ctaaggatcc gcaggcctct gctagcttga 481 ctgactgaga tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg 541 gacaaaccac aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta 601 ttgctttatt tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc 661 attttatgtt tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct 721 acaaatgtgg tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa 781 acgggtcttg aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg 841 cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag 901 caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt 961 atccttaagg tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt 1021 taatggtctt ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc 1081 cctaatctca ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact 1141 tggcgagatc gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat 1201 ctggtccttg ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa 1261 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc 1321 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga 1381 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc 1441 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt 1501 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct 1561 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg 1621 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta 1681 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct 1741 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa 1801 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt 1861 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta 1921 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat 1981 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa 2041 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct 2101 cagcgatctg tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc 2161 cctcgtcgga aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg 2221 aactatcgca agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg 2281 ccaggtatta ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct 2341 acatcctcaa tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc 2401 ctccgcgccg ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca 2461 cgtcagacga agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc 2521 ccgctgctca taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg 2581 ggacttgggt gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa 2641 gtagtccctt ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt 2701 atataaggac gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg 2761 gttcttgttt gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc 2821 ggggctttcg tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag 2881 ggctgtagtc tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc 2941 aaaatggcgg ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc 3001 gttgaaacaa ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct 3061 aatgcgggaa agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt 3121 gaagtttgtc actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg 3181 cggtgccgtt gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac 3241 gtcacccgtt ctgttggctt ataatgcagg gtggggccac cagccggtag gtgtgcggta 3301 ggcttttctc cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca 3361 ggcgccggac ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt 3421 atgtacctat cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc 3481 gagtgtgttt tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg 3541 taattttcag tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt 3601 gttagacaac atgggtaaaa agcctgaact caccgcgacg tctgtcgaga agtttctgat 3661 cgaaaagttc gacagcgtct ccgacctgat gcagctctcg gagggcgaag aatctcgtgc 3721 tttcagcttc gatgtaggag ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg 3781 tttctacaaa gatcgttatg tttatcggca ctttgcatcc gccgcgctcc cgattccgga 3841 agtgcttgac attggggagt tcagcgagag cctgacctat tgcatctccc gccgtgcaca 3901 gggtgtcacg ttgcaagacc tgcctgaaac cgaactgccc gctgttctgc agccggtcgc 3961 ggaggcaatg gatgccatcg ctgccgccga tcttagccag acgagcgggt tcggcccatt 4021 cggaccgcaa ggaatcggtc aatacactac atggcgtgat ttcatatgcg cgattgctga 4081 tccccatgtg tatcactggc aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca 4141 ggctctcgat gagctgatgc tttgggccga ggactgcccc gaagtccggc acctcgtgca 4201 cgcggatttc ggctccaaca atgtcctgac ggacaatggc cgcataacag cggtcattga 4261 ctggagcgag gcgatgttcg gggattccca atacgaggtc gccaacatct tcttctggag 4321 gccgtggttg gcttgtatgg agcagcagac gcgctacttc gagcggaggc atccggagct 4381 tgcaggatcg ccccggctcc gggcgtatat gctccgcatt ggtcttgacc aactctatca 4441 gagcttggtt gacggcaatt tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat 4501 cgtccgatcc ggagccggga ctgtcgggcg tacacaaatc gcccgcagaa gcgccgccgt 4561 ctggaccgat ggctgtgtag aagtactcgc cgatagtgga aaccgacgcc ccagcactcg 4621 tccgagggca aaggaataag ctagtatgta agcctagtct tagataataa aatcgctatc 4681 catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac gctaggcgcg cctggtgtac 4741 cgagaacgat cctctcagtg cgagtctcga cgatccatat cgttgcttgg cagtcagcca 4801 gtcggaatcc agcttgggac ccaggaagtc caatcgtcag atattgtact caagcctggt 4861 cacggcagcg taccgatctg tttaaaccta gatattgata gtctgatcgg tcaacgtata 4921 atcgagtcct agcttttgca aacatctatc aagagacagg atcagcagga ggctttcgca 4981 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg 5041 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcgc 5101 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg 5161 aagaacgctt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc 5221 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg 5281 ttgagtattc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat 5341 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgattg 5401 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg 5461 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc 5521 ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg cgaactactt actctagctt 5581 cccggcaaca gttgatagac tggatggagg cggataaagt tgcaggacca cttctgcgct 5641 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc 5701 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca 5761 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct 5821 cactgattaa gcattggtaa ccgattctag gtgcattggc gcagaaaaaa atgcctgatg 5881 cgacgctgcg cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc 5941 aacttgccca cttccatacg tgtcctcctt accagaaatt tatccttaag atcccgaatc 6001 gtttaaactc gactctggct ctatcgaatc tccgtcgttt cgagcttacg cgaacagccg 6061 tggcgctcat ttgctcgtcg ggcatcgaat ctcgtcagct atcgtcagct tacctttttg 6121 gcagcgatcg cggctcccga catcttggac cattagctcc acaggtatct tcttccctct 6181 agtggtcata acagcagctt cagctacctc tcaattcaaa aaacccctca agacccgttt 6241 agaggcccca aggggttatg ctatcaatcg ttgcgttaca cacacaaaaa accaacacac 6301 atccatcttc gatggatagc gattttatta tctaactgct gatcgagtgt agccagatct 6361 agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac 6421 ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa 6481 tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt 6541 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc 6601 ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat 6661 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atgctgatgc 6721 ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc 6781 tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa 6841 aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg 6901 tctatataag cagagctggt ttagtgaacc gtcagatcag atctttgtcg atcctaccat 6961 ccactcgaca cacccgccag cggccgc //