LOCUS       sequence-295705-        6748 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp8.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6748)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6748)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: pGBW-m4134427
FEATURES             Location/Qualifiers
     source          1..6748
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             602..622
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             623..664
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     polyA_signal    717..838
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(728..747)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     782..801
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      953..999
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1029..1165
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1210..1281
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1507..2095)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1587..1606)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2595..3804
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3641..3660
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3808..4407
                     /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(3808..3827)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4304..4324
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4434..4482
                     /note="synthetic polyadenylation signal"
     CDS             join(4748..4816,4817..5608)
                     /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(4966..4985)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5621..5757
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5902..5973
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5978..6024)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6042..6090
                     /note="synthetic polyadenylation signal"
     enhancer        6171..6474
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6476..6678
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6629..6649
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(6747..6748,1..8)
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 caccatggca attgcctcag aatttagttc cctcccctcg tacgctgcgt tcgctaccgc
       61 ccaagaggca tatgagcagg ccgtagccaa cggggacagc gaagtggttc ttaagaaact
      121 gaagaaatct ttgaatgtgg caaagagcga gtttgatcgg gacgcggcaa tgcagaggaa
      181 gctggaaaaa atggctgatc aggctatgac acagatgtac aaacaagccc gatccgagga
      241 caagagagct aaagtcactt ctgccatgca aacgatgcta ttcaccatgt tacgtaagct
      301 ggataacgac gcactcaata acataatcaa taacgcccgc gatggttgtg tgccattgaa
      361 tatcattcct cttactacag cggccaagct gatggtcgtt atcccggact ataacacata
      421 caaaaatacc tgcgacggca ccactttcac gtatgctagc gccctgtggg aaatacagca
      481 ggtagtggat gccgattcaa agattgtgca gcttagtgag atctccatgg acaacagtcc
      541 caacttggct tggcctttaa tcgtcacagc tctcagagcc aatagcgcag ttaaactgca
      601 agaaaacctt tactttcagt cggggaaacc tatacccaat ccacttctcg gcttagattc
      661 tacctaagct agcttgactg actgagatac agcgtacctt cagctcacag acatgataag
      721 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
      781 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
      841 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta
      901 aagcaagtaa aacctctaca aatgtggtat tggcccatct ctatcggtat cgtagcataa
      961 ccccttgggg cctctaaacg ggtcttgagg ggttttttgt gcccctcggg ccggattgct
     1021 atctaccggc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc
     1081 acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc catacgtgtc
     1141 ctccttacca gaaatttatc cttaaggtcg tcagctatcc tgcaggcgat ctctcgattt
     1201 cgatcaagac attcctttaa tggtcttttc tggacaccac taggggtcag aagtagttca
     1261 tcaaactttc ttccctccct aatctcattg gttaccttgg gctatcgaaa cttaattaac
     1321 cagtcaagtc agctacttgg cgagatcgac ttgtctgggt ttcgactacg ctcagaattg
     1381 cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt tctccgatta cgagtttcat
     1441 ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     1501 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     1561 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     1621 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     1681 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     1741 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     1801 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     1861 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     1921 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     1981 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     2041 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     2101 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     2161 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     2221 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     2281 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg catttaaatt
     2341 tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc tttcgtccga tccatcttgc
     2401 aggctacctc tcgaacgaac tatcgcaagt ctcttggccg gccttgcgcc ttggctattg
     2461 cttggcagcg cctatcgcca ggtattactc caatcccgaa tatccgagat cgggatcacc
     2521 cgagagaagt tcaacctaca tcctcaatcc cgatctatcc gagatccgag gaatatcgaa
     2581 atcggggcgc gcctggcctc cgcgccgggt tttggcgcct cccgcgggcg cccccctcgt
     2641 cacggcgagc gctgccacgt cagacgaagg gcgcaggagc gtcctgatcc ttccgcccgg
     2701 acgctcagga cagcggcccg ctgctcataa gactcggcct tagaacccca gtatcagcag
     2761 aaggacattt taggacggga cttgggtgac tctagggcac tggttttctt tccagagagc
     2821 ggaacaggcg aggaaaagta gtcccttctc ggcgattctg cggagggatc tccgtggggc
     2881 ggtgaacgcc gatgattata taaggacgcg ccgggtgtgg cacagctagt tccgtcgcag
     2941 ccgggatttg ggtcgcggtt cttgtttgtg gatcgctgtg atcgtcactt ggtgagtagc
     3001 gggctgctgg gctggccggg gctttcgtgg ccgccgggcc gctcggtggg acggaagcgt
     3061 gtggagagac cgccaagggc tgtagtctgg gtccgcgagc aaggttgccc tgaactgggg
     3121 gttgggggga gcgcagcaaa atggcggctg ttcccgagtc ttgaatggaa gacgcttgtg
     3181 aggcgggctg tgaggtcgtt gaaacaaggt ggggggcatg gtgggcggca agaacccaag
     3241 gtcttgagcc cttcgctaat gcgggaaagc tcttattcgg gtgagatggg ctgggcacca
     3301 tctggggacc ctgacgtgaa gtttgtcact gactggagaa ctcggtttgt cgtctgttgc
     3361 gggggcggca gttatggcgg tgccgttggg cagtgcaccc gtacctttgg gagcgcgcgc
     3421 cctcgtcgtg tcgtgacgtc acccgttctg ttggcttata atgcagggtg gggccacctg
     3481 ccggtaggtg tgcggtaggc ttttctccgt cgcaggacgc agggttcggg cctagggtag
     3541 gctctcctga atcgacaggc gccggacctc tggtgagggg agggataagt gaggcgtcag
     3601 tttctttggt cggttttatg tacctatctt cttaagtagc tgaagctccg gttttgaact
     3661 atgcgctcgg ggttggcgag tgtgttttgt gaagtttttt aggcaccttt tgaaatgtaa
     3721 tcatttgggt caatatgtaa ttttcagtgt tagacttgta aattgtccgc taaattctgg
     3781 ccgtttttgg cttttttgtt agacaacatg accgagtaca agcccacggt gcgcctcgcc
     3841 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     3901 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     3961 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     4021 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     4081 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggagggc
     4141 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggcgtctcg
     4201 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggccgcc
     4261 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     4321 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     4381 tgcatgaccc gcaagcccgg tgcctaggct agtatgtaag cctagtctta gataataaaa
     4441 tcgctatcca tcgaagatgg atgtgtgttg gttttttgtg tgtgtaacgc taggcgcgcc
     4501 tggtgtaccg agaacgatcc tctcagtgcg agtctcgacg atccatatcg ttgcttggca
     4561 gtcagccagt cggaatccag cttgggaccc aggaagtcca atcgtcagat attgtactca
     4621 agcctggtca cggcagcgta ccgatctgtt taaacctaga tattgatagt ctgatcggtc
     4681 aacgtataat cgagtcctag cttttgcaaa catctatcaa gagacaggat cagcaggagg
     4741 ctttcgcatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     4801 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     4861 gggtgcgcga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     4921 tcgccccgaa gaacgctttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     4981 attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     5041 tgacttggtt gagtattcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     5101 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     5161 aacgattgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     5221 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     5281 cacgatgcct gtagcaatgg caacaacctt gcgtaaacta ttaactggcg aactacttac
     5341 tctagcttcc cggcaacagt tgatagactg gatggaggcg gataaagttg caggaccact
     5401 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     5461 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     5521 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     5581 aggtgcctca ctgattaagc attggtaacc gattctaggt gcattggcgc agaaaaaaat
     5641 gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc aacggatacg
     5701 gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta tccttaagat
     5761 cgtttaaact cgactctggc tctatcgaat ctccgtcgtt tcgagcttac gcgaacagcc
     5821 gtggcgctca tttgctcgtc gggcatcgaa tctcgtcagc tatcgtcagc ttaccttttt
     5881 ggcagcgatc gcggctcccg acatcttgga ccattagctc cacaggtatc ttcttccctc
     5941 tagtggtcat aacagcagct tcagctacct ctcaattcaa aaaacccctc aagacccgtt
     6001 tagaggcccc aaggggttat gctatcaatc gttgcgttac acacacaaaa aaccaacaca
     6061 catccatctt cgatggatag cgattttatt atctaactgc tgatcgagtg tagccagatc
     6121 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
     6181 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
     6241 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
     6301 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
     6361 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
     6421 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catgctgatg
     6481 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
     6541 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
     6601 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
     6661 gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc gatcctacca
     6721 tccactcgac acacccgcca gcggccgc
//
