LOCUS Exported 5103 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Mammalian expression plasmid for SARS-CoV-2 ORF3a protein. ACCESSION . VERSION . KEYWORDS pGBW-m4133902 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 5103) TITLE SARS-CoV-2 protein Mammalian expression constructs, tagged, for protein expression and/or purification REFERENCE 2 (bases 1 to 5103) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..5103 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 75..378 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 380..582 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 533..553 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" regulatory 651..660 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 1482..1502 /codon_start=1 /product="tobacco etch virus (TEV) protease recognition and cleavage site" /label=TEV site /translation="ENLYFQS" CDS 1506..1571 /codon_start=1 /product="three tandem FLAG(R) epitope tags, followed by an enterokinase cleavage site" /label=3xFLAG /translation="DYKDHDGDYKDHDIDYKDDDDK" polyA_signal 1624..1745 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(1635..1654) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 1689..1708 /label=SV40pA-R /note="SV40 polyA, reverse primer" terminator 1860..1906 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" terminator 1936..2072 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 2117..2188 /label=3' beta-globin insulator /note="insulator downstream of the human beta-globin locus (Farrell et al., 2002)" rep_origin complement(2414..3002) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(2494..2513) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS 3748..4608 /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(3966..3985) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" terminator 4621..4757 /label=rrnG terminator /note="transcription terminator from the E. coli ribosomal RNA rrnG operon (Albrechtsen et al., 1991)" misc_feature 4909..4980 /label=5' beta-globin insulator /note="insulator upstream of the human beta-globin locus (Farrell et al., 2002)" terminator 4985..5031 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" polyA_signal 5049..5097 /note="synthetic polyadenylation signal" ORIGIN 1 ctgctgatcg agtgtagcca gatctagtaa tcaattacgg ggtcattagt tcatagccca 61 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac 121 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact 181 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa 241 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg 301 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta 361 gtcatcgcta ttaccatgct gatgcggttt tggcagtaca tcaatgggcg tggatagcgg 421 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg 481 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg 541 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctggtttagt gaaccgtcag 601 atcagatctt tgtcgatcct accatccact cgacacaccc gccagcggcc gccaccatgg 661 atttgtttat gagaatcttc acaattggaa ctgtaacttt gaagcaaggt gaaatcaagg 721 atgctactcc ttcagatttt gttcgcgcta ctgcaacgat accgatacaa gcctcactcc 781 ctttcggatg gcttattgtt ggcgttgcac ttcttgctgt ttttcagagc gcttccaaaa 841 tcataaccct caaaaagaga tggcaactag cactctccaa gggtgttcac tttgtttgca 901 acttgctgtt gttgtttgta acagtttact cacacctttt gctcgttgct gctggccttg 961 aagccccttt tctctatctt tatgctttag tctacttctt gcagagtata aactttgtaa 1021 gaataataat gaggctttgg ctttgctgga aatgccgttc caaaaaccca ttactttatg 1081 atgccaacta ttttctttgc tggcatacta attgttacga ctattgtata ccttacaata 1141 gtgtaacttc ttcaattgtc attacttcag gtgatggcac aacaagtcct atttctgaac 1201 atgactacca gattggtggt tatactgaaa aatgggaatc tggagtaaaa gactgtgttg 1261 tattacacag ttacttcact tcagactatt accagctgta ctcaactcaa ttgagtacag 1321 acactggtgt tgaacatgtt accttcttca tctacaataa aattgttgat gagcctgaag 1381 aacatgtcca aattcacaca atcgacggtt catccggagt tgttaatcca gtaatggaac 1441 caatttatga tgaaccgacg acgactacta gcgtgccttt ggaaaacctt tactttcagt 1501 cgatggatta taaggaccac gatggagact acaaagatca tgacattgat tacaaggacg 1561 acgatgacaa ataagctagc ttgactgact gagatacagc gtaccttcag ctcacagaca 1621 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct 1681 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac 1741 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg 1801 ttttttaaag caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt 1861 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg 1921 gattgctatc taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta 1981 tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat 2041 acgtgtcctc cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc 2101 tcgatttcga tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag 2161 tagttcatca aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt 2221 aattaaccag tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc 2281 agaattgcgt cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga 2341 gtttcattta aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg 2401 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct 2461 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa 2521 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 2581 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 2641 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 2701 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 2761 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 2821 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc 2881 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 2941 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 3001 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 3061 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa 3121 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat 3181 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat 3241 ttaaatttcc gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc 3301 atcttgcagg ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg 3361 gctattgctt ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg 3421 gatcacccga gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa 3481 tatcgaaatc ggggcgcgcc tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg 3541 atccatatcg ttgcttggca gtcagccagt cggaatccag cttgggaccc aggaagtcca 3601 atcgtcagat attgtactca agcctggtca cggcagcgta ccgatctgtt taaacctaga 3661 tattgatagt ctgatcggtc aacgtataat cgagtcctag cttttgcaaa catctatcaa 3721 gagacaggat cagcaggagg ctttcgcatg agtattcaac atttccgtgt cgcccttatt 3781 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta 3841 aaagatgctg aagatcagtt gggtgcgcga gtgggttaca tcgaactgga tctcaacagc 3901 ggtaagatcc ttgagagttt tcgccccgaa gaacgctttc caatgatgag cacttttaaa 3961 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc 4021 cgcatacact attctcagaa tgacttggtt gagtattcac cagtcacaga aaagcatctt 4081 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact 4141 gcggccaact tacttctgac aacgattgga ggaccgaagg agctaaccgc ttttttgcac 4201 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata 4261 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacctt gcgtaaacta 4321 ttaactggcg aactacttac tctagcttcc cggcaacagt tgatagactg gatggaggcg 4381 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat 4441 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt 4501 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga 4561 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaacc gattctaggt 4621 gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc 4681 cagattcagc aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac 4741 cagaaattta tccttaagat cccgaatcgt ttaaactcga ctctggctct atcgaatctc 4801 cgtcgtttcg agcttacgcg aacagccgtg gcgctcattt gctcgtcggg catcgaatct 4861 cgtcagctat cgtcagctta cctttttggc agcgatcgcg gctcccgaca tcttggacca 4921 ttagctccac aggtatcttc ttccctctag tggtcataac agcagcttca gctacctctc 4981 aattcaaaaa acccctcaag acccgtttag aggccccaag gggttatgct atcaatcgtt 5041 gcgttacaca cacaaaaaac caacacacat ccatcttcga tggatagcga ttttattatc 5101 taa //