LOCUS Exported 6170 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp11. ACCESSION . VERSION . KEYWORDS pGBW-m4252796 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6170) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6170) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6170 /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 62..79 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 132..253 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(143..162) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 197..216 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 368..414 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 444..580 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 625..696 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(922..1510) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1002..1021) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 2010..3219 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3056..3075 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 3223..3822 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(3223..3242) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 3719..3739 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 3849..3897 /note="synthetic polyadenylation signal" CDS 4163..5023 /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(4381..4400) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5036..5172 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 5324..5395 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 5400..5446 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5464..5512 /note="synthetic polyadenylation signal" enhancer 5593..5896 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 5898..6100 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6051..6071 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 6169..8 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" ORIGIN 1 caccatggct gatgcacaat cgtttttaaa cgggtttgcg gaaaaccttt actttcagtc 61 gcaccatcac catcaccatt aagctagctt gactgactga gatacagcgt accttcagct 121 cacagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa 181 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg 241 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt 301 gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtattggcc catctctatc 361 ggtatcgtag cataacccct tggggcctct aaacgggtct tgaggggttt tttgtgcccc 421 tcgggccgga ttgctatcta ccggcattgg cgcagaaaaa aatgcctgat gcgacgctgc 481 gcgtcttata ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc 541 acttccatac gtgtcctcct taccagaaat ttatccttaa ggtcgtcagc tatcctgcag 601 gcgatctctc gatttcgatc aagacattcc tttaatggtc ttttctggac accactaggg 661 gtcagaagta gttcatcaaa ctttcttccc tccctaatct cattggttac cttgggctat 721 cgaaacttaa ttaaccagtc aagtcagcta cttggcgaga tcgacttgtc tgggtttcga 781 ctacgctcag aattgcgtca gtcaagttcg atctggtcct tgctattgca cccgttctcc 841 gattacgagt ttcatttaaa tcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa 901 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa 961 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc 1021 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc 1081 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag 1141 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga 1201 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc 1261 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac 1321 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg 1381 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca 1441 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa 1501 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa 1561 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt 1621 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag 1681 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat 1741 agttgcattt aaatttccga actctccaag gccctcgtcg gaaaatcttc aaacctttcg 1801 tccgatccat cttgcaggct acctctcgaa cgaactatcg caagtctctt ggccggcctt 1861 gcgccttggc tattgcttgg cagcgcctat cgccaggtat tactccaatc ccgaatatcc 1921 gagatcggga tcacccgaga gaagttcaac ctacatcctc aatcccgatc tatccgagat 1981 ccgaggaata tcgaaatcgg ggcgcgcctg gcctccgcgc cgggttttgg cgcctcccgc 2041 gggcgccccc ctcgtcacgg cgagcgctgc cacgtcagac gaagggcgca ggagcgtcct 2101 gatccttccg cccggacgct caggacagcg gcccgctgct cataagactc ggccttagaa 2161 ccccagtatc agcagaagga cattttagga cgggacttgg gtgactctag ggcactggtt 2221 ttctttccag agagcggaac aggcgaggaa aagtagtccc ttctcggcga ttctgcggag 2281 ggatctccgt ggggcggtga acgccgatga ttatataagg acgcgccggg tgtggcacag 2341 ctagttccgt cgcagccggg atttgggtcg cggttcttgt ttgtggatcg ctgtgatcgt 2401 cacttggtga gtagcgggct gctgggctgg ccggggcttt cgtggccgcc gggccgctcg 2461 gtgggacgga agcgtgtgga gagaccgcca agggctgtag tctgggtccg cgagcaaggt 2521 tgccctgaac tgggggttgg ggggagcgca gcaaaatggc ggctgttccc gagtcttgaa 2581 tggaagacgc ttgtgaggcg ggctgtgagg tcgttgaaac aaggtggggg gcatggtggg 2641 cggcaagaac ccaaggtctt gagcccttcg ctaatgcggg aaagctctta ttcgggtgag 2701 atgggctggg caccatctgg ggaccctgac gtgaagtttg tcactgactg gagaactcgg 2761 tttgtcgtct gttgcggggg cggcagttat ggcggtgccg ttgggcagtg cacccgtacc 2821 tttgggagcg cgcgccctcg tcgtgtcgtg acgtcacccg ttctgttggc ttataatgca 2881 gggtggggcc accagccggt aggtgtgcgg taggcttttc tccgtcgcag gacgcagggt 2941 tcgggcctag ggtaggctct cctgaatcga caggcgccgg acctctggtg aggggaggga 3001 taagtgaggc gtcagtttct ttggtcggtt ttatgtacct atcttcttaa gtagctgaag 3061 ctccggtttt gaactatgcg ctcggggttg gcgagtgtgt tttgtgaagt tttttaggca 3121 ccttttgaaa tgtaatcatt tgggtcaata tgtaattttc agtgttagac ttgtaaattg 3181 tccgctaaat tctggccgtt tttggctttt ttgttagaca acatgaccga gtacaagccc 3241 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg 3301 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc 3361 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc 3421 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg 3481 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag 3541 caacagatgg agggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc 3601 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc 3661 ggagtggagg ccgccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc 3721 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa 3781 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct aggctagtat gtaagcctag 3841 tcttagataa taaaatcgct atccatcgaa gatggatgtg tgttggtttt ttgtgtgtgt 3901 aacgctaggc gcgcctggtg taccgagaac gatcctctca gtgcgagtct cgacgatcca 3961 tatcgttgct tggcagtcag ccagtcggaa tccagcttgg gacccaggaa gtccaatcgt 4021 cagatattgt actcaagcct ggtcacggca gcgtaccgat ctgtttaaac ctagatattg 4081 atagtctgat cggtcaacgt ataatcgagt cctagctttt gcaaacatct atcaagagac 4141 aggatcagca ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc ttattccctt 4201 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga 4261 tgctgaagat cagttgggtg cgcgagtggg ttacatcgaa ctggatctca acagcggtaa 4321 gatccttgag agttttcgcc ccgaagaacg ctttccaatg atgagcactt ttaaagttct 4381 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat 4441 acactattct cagaatgact tggttgagta ttcaccagtc acagaaaagc atcttacgga 4501 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc 4561 caacttactt ctgacaacga ttggaggacc gaaggagcta accgcttttt tgcacaacat 4621 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa 4681 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca accttgcgta aactattaac 4741 tggcgaacta cttactctag cttcccggca acagttgata gactggatgg aggcggataa 4801 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc 4861 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc 4921 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag 4981 acagatcgct gagataggtg cctcactgat taagcattgg taaccgattc taggtgcatt 5041 ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca tatgccagat 5101 tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc cttaccagaa 5161 atttatcctt aagatcccga atcgtttaaa ctcgactctg gctctatcga atctccgtcg 5221 tttcgagctt acgcgaacag ccgtggcgct catttgctcg tcgggcatcg aatctcgtca 5281 gctatcgtca gcttaccttt ttggcagcga tcgcggctcc cgacatcttg gaccattagc 5341 tccacaggta tcttcttccc tctagtggtc ataacagcag cttcagctac ctctcaattc 5401 aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctatcaa tcgttgcgtt 5461 acacacacaa aaaaccaaca cacatccatc ttcgatggat agcgatttta ttatctaact 5521 gctgatcgag tgtagccaga tctagtaatc aattacgggg tcattagttc atagcccata 5581 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga 5641 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt 5701 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt 5761 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca 5821 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt 5881 catcgctatt accatgctga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt 5941 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca 6001 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg 6061 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat 6121 cagatctttg tcgatcctac catccactcg acacacccgc cagcggccgc //