LOCUS Exported 4845 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 nsp8. ACCESSION . VERSION . KEYWORDS pGBW-m4133147 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4845) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 4845) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4845 /organism="synthetic DNA construct" /mol_type="other DNA" CDS 596..616 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 617..661 /codon_start=1 /product="affinity and epitope tag derived from pancreatic ribonuclease A" /label=S-Tag /translation="KETAAAKFERQHMDS" polyA_signal 714..835 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(725..744) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 779..798 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 950..996 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1026..1162 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 1207..1278 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(1504..2092) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(1584..1603) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 2838..3698 /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(3056..3075) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 3711..3847 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 3999..4070 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4075..4121 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 4139..4187 /note="synthetic polyadenylation signal" enhancer 4268..4571 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 4573..4775 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 4726..4746 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" ORIGIN 1 caccatgata gcctcagagt ttagttccct tccatcatat gcagcttttg ctactgctca 61 agaagcttat gagcaggctg ttgctaatgg tgattctgaa gttgttctta aaaagttgaa 121 gaagtctttg aatgtggcta aatctgaatt tgaccgtgat gcagccatgc aacgtaagtt 181 ggaaaagatg gctgatcaag ctatgaccca aatgtataaa caggctagat ctgaggacaa 241 gagggcaaaa gttactagtg ctatgcagac aatgcttttc actatgctta gaaagttgga 301 taatgatgca ctcaacaaca ttatcaacaa tgcaagagat ggttgtgttc ccttgaacat 361 aatacctctt acaacagcag ccaaactaat ggttgtcata ccagactata acacatataa 421 aaatacgtgt gatggtacaa catttactta tgcatcagca ttgtgggaaa tccaacaggt 481 tgtagatgca gatagtaaaa ttgttcaact tagtgaaatt agtatggaca attcacctaa 541 tttagcatgg cctcttattg taacagcttt aagggccaat tctgctgtca aattagaaaa 601 cctttacttt cagtcgaaag aaaccgcggc agctaagttc gagaggcagc acatggacag 661 ttaagctagc ttgactgact gagatacagc gtaccttcag ctcacagaca tgataagata 721 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga 781 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa 841 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag 901 caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt agcataaccc 961 cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg gattgctatc 1021 taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca 1081 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc 1141 cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc tcgatttcga 1201 tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag tagttcatca 1261 aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt aattaaccag 1321 tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc agaattgcgt 1381 cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga gtttcattta 1441 aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc 1501 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag 1561 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt 1621 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg 1681 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg 1741 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg 1801 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac 1861 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg 1921 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt 1981 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg 2041 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc 2101 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt 2161 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt 2221 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag 2281 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat ttaaatttcc 2341 gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc atcttgcagg 2401 ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg gctattgctt 2461 ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg gatcacccga 2521 gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa tatcgaaatc 2581 ggggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg 2641 ttgcttggca gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat 2701 attgtactca agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt 2761 ctgatcggtc aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat 2821 cagcaggagg ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg 2881 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg 2941 aagatcagtt gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc 3001 ttgagagttt tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat 3061 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact 3121 attctcagaa tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca 3181 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact 3241 tacttctgac aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg 3301 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg 3361 agcgtgacac cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg 3421 aactacttac tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg 3481 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag 3541 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc 3601 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga 3661 tcgctgagat aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc 3721 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc 3781 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta 3841 tccttaagat cccgaatcgt ttaaactcga ctctggctct atcgaatctc cgtcgtttcg 3901 agcttacgcg aacagccgtg gcgctcattt gctcgtcggg catcgaatct cgtcagctat 3961 cgtcagctta cctttttggc agcgatcgcg gctcccgaca tcttggacca ttagctccac 4021 aggtatcttc ttccctctag tggtcataac agcagcttca gctacctctc aattcaaaaa 4081 acccctcaag acccgtttag aggccccaag gggttatgct atcaatcgtt gcgttacaca 4141 cacaaaaaac caacacacat ccatcttcga tggatagcga ttttattatc taactgctga 4201 tcgagtgtag ccagatctag taatcaatta cggggtcatt agttcatagc ccatatatgg 4261 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc 4321 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt 4381 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc 4441 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg 4501 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg 4561 ctattaccat gctgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact 4621 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa 4681 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta 4741 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatcagat 4801 ctttgtcgat cctaccatcc actcgacaca cccgccagcg gccgc //