LOCUS Exported 6548 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp11. ACCESSION . VERSION . KEYWORDS pGBW-m4252816 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6548) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 6548) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6548 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 47..67 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 416..433 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" polyA_signal 510..631 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(521..540) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 575..594 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 746..792 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 822..958 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1003..1074 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1300..1888) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1380..1399) /label=pBR322ori-F /note="pBR322 origin, forward primer" promoter 2388..3597 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3434..3453 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 3601..4200 /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(3601..3620) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 4097..4117 /label=Puro-F /note="Puromycin resistance gene, forward primer" polyA_signal 4227..4275 /note="synthetic polyadenylation signal" CDS 4541..5401 /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(4759..4778) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 5414..5550 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 5702..5773 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 5778..5824 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5842..5890 /note="synthetic polyadenylation signal" enhancer 5971..6274 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 6276..6478 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 6429..6449 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgagt gccgatgctc agtcctttct caacggtttc gcagtggaaa acctttactt 61 tcagtcgatg agtgacaaaa tcattcatct aaccgatgac tcctttgata cagacgtgtt 121 gaaggccgat ggtgcgatac tggtagattt ctgggctgaa tggtgtggcc cgtgcaagat 181 gatcgcaccc attctggacg agatcgccga cgagtatcaa ggaaaattaa ctgtcgctaa 241 gcttaatata gatcagaacc cagggacggc acctaaatac ggcatcaggg gaatacctac 301 cctcctgctc tttaagaatg gggaagttgc cgcgacaaaa gtgggagccc tgtctaaggg 361 ccagctaaaa gagttcttgg acgctaacct ggcaggttca ggctcggggc acatgcacca 421 tcaccatcac catagcagcg ggctcgtgcc acgcggctaa gctagcttga ctgactgaga 481 tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg gacaaaccac 541 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt 601 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt 661 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg 721 tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa acgggtcttg 781 aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg cagaaaaaaa 841 tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac 901 ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg 961 tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt taatggtctt 1021 ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc cctaatctca 1081 ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact tggcgagatc 1141 gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat ctggtccttg 1201 ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa aaggccagca 1261 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 1321 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 1381 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 1441 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 1501 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 1561 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 1621 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 1681 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 1741 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 1801 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 1861 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 1921 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 1981 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga 2041 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg 2101 tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc cctcgtcgga 2161 aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg aactatcgca 2221 agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg ccaggtatta 2281 ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct acatcctcaa 2341 tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggc ctccgcgccg 2401 ggttttggcg cctcccgcgg gcgcccccct cgtcacggcg agcgctgcca cgtcagacga 2461 agggcgcagg agcgtcctga tccttccgcc cggacgctca ggacagcggc ccgctgctca 2521 taagactcgg ccttagaacc ccagtatcag cagaaggaca ttttaggacg ggacttgggt 2581 gactctaggg cactggtttt ctttccagag agcggaacag gcgaggaaaa gtagtccctt 2641 ctcggcgatt ctgcggaggg atctccgtgg ggcggtgaac gccgatgatt atataaggac 2701 gcgccgggtg tggcacagct agttccgtcg cagccgggat ttgggtcgcg gttcttgttt 2761 gtggatcgct gtgatcgtca cttggtgagt agcgggctgc tgggctggcc ggggctttcg 2821 tggccgccgg gccgctcggt gggacggaag cgtgtggaga gaccgccaag ggctgtagtc 2881 tgggtccgcg agcaaggttg ccctgaactg ggggttgggg ggagcgcagc aaaatggcgg 2941 ctgttcccga gtcttgaatg gaagacgctt gtgaggcggg ctgtgaggtc gttgaaacaa 3001 ggtggggggc atggtgggcg gcaagaaccc aaggtcttga gcccttcgct aatgcgggaa 3061 agctcttatt cgggtgagat gggctgggca ccatctgggg accctgacgt gaagtttgtc 3121 actgactgga gaactcggtt tgtcgtctgt tgcgggggcg gcagttatgg cggtgccgtt 3181 gggcagtgca cccgtacctt tgggagcgcg cgccctcgtc gtgtcgtgac gtcacccgtt 3241 ctgttggctt ataatgcagg gtggggccac cagccggtag gtgtgcggta ggcttttctc 3301 cgtcgcagga cgcagggttc gggcctaggg taggctctcc tgaatcgaca ggcgccggac 3361 ctctggtgag gggagggata agtgaggcgt cagtttcttt ggtcggtttt atgtacctat 3421 cttcttaagt agctgaagct ccggttttga actatgcgct cggggttggc gagtgtgttt 3481 tgtgaagttt tttaggcacc ttttgaaatg taatcatttg ggtcaatatg taattttcag 3541 tgttagactt gtaaattgtc cgctaaattc tggccgtttt tggctttttt gttagacaac 3601 atgaccgagt acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta 3661 cgcaccctcg ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac 3721 cgccacatcg agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac 3781 atcggcaagg tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag 3841 agcgtcgaag cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt 3901 tcccggctgg ccgcgcagca acagatggag ggcctcctgg cgccgcaccg gcccaaggag 3961 cccgcgtggt tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc 4021 agcgccgtcg tgctccccgg agtggaggcc gccgagcgcg ccggggtgcc cgccttcctg 4081 gagacctccg cgccccgcaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc 4141 gacgtcgagg tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctag 4201 gctagtatgt aagcctagtc ttagataata aaatcgctat ccatcgaaga tggatgtgtg 4261 ttggtttttt gtgtgtgtaa cgctaggcgc gcctggtgta ccgagaacga tcctctcagt 4321 gcgagtctcg acgatccata tcgttgcttg gcagtcagcc agtcggaatc cagcttggga 4381 cccaggaagt ccaatcgtca gatattgtac tcaagcctgg tcacggcagc gtaccgatct 4441 gtttaaacct agatattgat agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc 4501 aaacatctat caagagacag gatcagcagg aggctttcgc atgagtattc aacatttccg 4561 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac 4621 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgcg cgagtgggtt acatcgaact 4681 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgct ttccaatgat 4741 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga 4801 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac 4861 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat 4921 gagtgataac actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac 4981 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct 5041 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac 5101 cttgcgtaaa ctattaactg gcgaactact tactctagct tcccggcaac agttgataga 5161 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg 5221 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact 5281 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac 5341 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta 5401 accgattcta ggtgcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata 5461 ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc acttccatac 5521 gtgtcctcct taccagaaat ttatccttaa gatcccgaat cgtttaaact cgactctggc 5581 tctatcgaat ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc 5641 gggcatcgaa tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg 5701 acatcttgga ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct 5761 tcagctacct ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat 5821 gctatcaatc gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag 5881 cgattttatt atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc 5941 attagttcat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc 6001 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt 6061 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca 6121 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg 6181 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca 6241 gtacatctac gtattagtca tcgctattac catgctgatg cggttttggc agtacatcaa 6301 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa 6361 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc 6421 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgg 6481 tttagtgaac cgtcagatca gatctttgtc gatcctacca tccactcgac acacccgcca 6541 gcggccgc //