LOCUS Exported 5319 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 endoRNAse. ACCESSION . VERSION . KEYWORDS pGBW-m4134159 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5319) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5319) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5319 /organism="synthetic DNA construct" /mol_type="other DNA" terminator 228..364 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 516..587 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 592..638 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 656..704 /note="synthetic polyadenylation signal" enhancer 785..1088 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 1090..1292 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 1243..1263 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" CDS 2408..2428 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 2432..2497 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" polyA_signal 2550..2671 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(2561..2580) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 2615..2634 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 2786..2832 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 2862..2998 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3043..3114 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(3340..3928) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(3420..3439) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 4674..215 /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(4892..4911) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" ORIGIN 1 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg 61 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta 121 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg 181 ctgagatagg tgcctcactg attaagcatt ggtaaccgat tctaggtgca ttggcgcaga 241 aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca catatgccag attcagcaac 301 ggatacggct tccccaactt gcccacttcc atacgtgtcc tccttaccag aaatttatcc 361 ttaagatccc gaatcgttta aactcgactc tggctctatc gaatctccgt cgtttcgagc 421 ttacgcgaac agccgtggcg ctcatttgct cgtcgggcat cgaatctcgt cagctatcgt 481 cagcttacct ttttggcagc gatcgcggct cccgacatct tggaccatta gctccacagg 541 tatcttcttc cctctagtgg tcataacagc agcttcagct acctctcaat tcaaaaaacc 601 cctcaagacc cgtttagagg ccccaagggg ttatgctatc aatcgttgcg ttacacacac 661 aaaaaaccaa cacacatcca tcttcgatgg atagcgattt tattatctaa ctgctgatcg 721 agtgtagcca gatctagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt 781 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc 841 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac 901 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata 961 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc 1021 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta 1081 ttaccatgct gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac 1141 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc 1201 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc 1261 gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag atcagatctt 1321 tgtcgatcct accatccact cgacacaccc gccagcggcc gccaccatgt cactggagaa 1381 cgttgctttc aatgtagtga acaaggggca ctttgacgga cagcaaggcg aagtgccggt 1441 cagtatcata aataatacgg tgtacaccaa agtcgatggt gttgacgtgg agttatttga 1501 aaacaagaca actttgcctg tcaatgtggc cttcgagctg tgggcaaaac gcaacattaa 1561 gccagtaccc gaagtgaaaa tcctcaacaa tcttggcgtt gatattgccg caaacactgt 1621 tatctgggat tataagcgag atgcgcctgc ccatatatcc accattggag tgtgttctat 1681 gacagacatc gctaaaaagc ccaccgagac aatttgcgcg ccactcacgg tattctttga 1741 cgggagggtc gacggtcagg tggatctgtt tagaaatgcc cgtaacggcg tacttattac 1801 tgaaggaagc gtgaaaggcc tacagcccag tgttgggccg aagcaagcaa gcctaaatgg 1861 agtcacactg atcggagagg ctgtgaaaac ccagttcaac tactataaga aggtcgacgg 1921 cgtggttcag cagctgcctg agacatactt tacccaatcg cggaatctgc aagaatttaa 1981 gcccagatcc cagatggaga ttgacttctt ggaactggcc atggatgagt tcatcgaacg 2041 gtataagctc gagggctacg ccttcgagca catagtttat ggggattttt ctcattcaca 2101 gttaggcggt ctgcacttgc tcatcggatt agcgaaacgc tttaaagaga gcccattcga 2161 acttgaagac tttatcccaa tggactcaac tgtgaagaat tacttcataa ctgacgctca 2221 gacagggtcc agcaagtgcg tctgttctgt tattgatttg ctacttgatg atttcgtgga 2281 gattattaaa tcacaggacc tgtccgtggt ttcgaaggtc gtgaaagtaa ccatcgacta 2341 tacggaaatt agtttcatgc tttggtgtaa ggatggccat gtggagacat tttatcctaa 2401 gctccaggaa aacctttact ttcagtcgat ggattataag gaccacgatg gagactacaa 2461 agatcatgac attgattaca aggacgacga tgacaaataa gctagcttga ctgactgaga 2521 tacagcgtac cttcagctca cagacatgat aagatacatt gatgagtttg gacaaaccac 2581 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt 2641 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt 2701 tcaggttcag ggggaggtgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg 2761 tattggccca tctctatcgg tatcgtagca taaccccttg gggcctctaa acgggtcttg 2821 aggggttttt tgtgcccctc gggccggatt gctatctacc ggcattggcg cagaaaaaaa 2881 tgcctgatgc gacgctgcgc gtcttatact cccacatatg ccagattcag caacggatac 2941 ggcttcccca acttgcccac ttccatacgt gtcctcctta ccagaaattt atccttaagg 3001 tcgtcagcta tcctgcaggc gatctctcga tttcgatcaa gacattcctt taatggtctt 3061 ttctggacac cactaggggt cagaagtagt tcatcaaact ttcttccctc cctaatctca 3121 ttggttacct tgggctatcg aaacttaatt aaccagtcaa gtcagctact tggcgagatc 3181 gacttgtctg ggtttcgact acgctcagaa ttgcgtcagt caagttcgat ctggtccttg 3241 ctattgcacc cgttctccga ttacgagttt catttaaatc atgtgagcaa aaggccagca 3301 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 3361 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 3421 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 3481 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 3541 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 3601 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 3661 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 3721 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 3781 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 3841 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 3901 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 3961 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 4021 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga 4081 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg 4141 tctatttcgt tcatccatag ttgcatttaa atttccgaac tctccaaggc cctcgtcgga 4201 aaatcttcaa acctttcgtc cgatccatct tgcaggctac ctctcgaacg aactatcgca 4261 agtctcttgg ccggccttgc gccttggcta ttgcttggca gcgcctatcg ccaggtatta 4321 ctccaatccc gaatatccga gatcgggatc acccgagaga agttcaacct acatcctcaa 4381 tcccgatcta tccgagatcc gaggaatatc gaaatcgggg cgcgcctggt gtaccgagaa 4441 cgatcctctc agtgcgagtc tcgacgatcc atatcgttgc ttggcagtca gccagtcgga 4501 atccagcttg ggacccagga agtccaatcg tcagatattg tactcaagcc tggtcacggc 4561 agcgtaccga tctgtttaaa cctagatatt gatagtctga tcggtcaacg tataatcgag 4621 tcctagcttt tgcaaacatc tatcaagaga caggatcagc aggaggcttt cgcatgagta 4681 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg 4741 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcgcgagtgg 4801 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac 4861 gctttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg 4921 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt 4981 attcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg 5041 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg attggaggac 5101 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt 5161 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag 5221 caatggcaac aaccttgcgt aaactattaa ctggcgaact acttactcta gcttcccggc 5281 aacagttgat agactggatg gaggcggata aagttgcag //