LOCUS       sequence-320778-        6776 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 6776)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6776)
  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-m4134121
FEATURES             Location/Qualifiers
     source          1..6776
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1174..1194
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1198..1263
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    1316..1437
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1327..1346)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1381..1400
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1552..1598
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1628..1764
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1809..1880
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2106..2694)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2186..2205)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3194..4403
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4240..4259
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4407..5006
                     /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(4407..4426)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4903..4923
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5033..5081
                     /note="synthetic polyadenylation signal"
     CDS             join(5347..5415,5416..6207)
                     /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(5565..5584)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6220..6356
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6508..6579
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6584..6630)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6648..6696
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc
      541 gatcctacca tccactcgac acacccgcca gcggccgcca ccatgatagc ctcagagttt
      601 agttcccttc catcatatgc agcttttgct actgctcaag aagcttatga gcaggctgtt
      661 gctaatggtg attctgaagt tgttcttaaa aagttgaaga agtctttgaa tgtggctaaa
      721 tctgaatttg accgtgatgc agccatgcaa cgtaagttgg aaaagatggc tgatcaagct
      781 atgacccaaa tgtataaaca ggctagatct gaggacaaga gggcaaaagt tactagtgct
      841 atgcagacaa tgcttttcac tatgcttaga aagttggata atgatgcact caacaacatt
      901 atcaacaatg caagagatgg ttgtgttccc ttgaacataa tacctcttac aacagcagcc
      961 aaactaatgg ttgtcatacc agactataac acatataaaa atacgtgtga tggtacaaca
     1021 tttacttatg catcagcatt gtgggaaatc caacaggttg tagatgcaga tagtaaaatt
     1081 gttcaactta gtgaaattag tatggacaat tcacctaatt tagcatggcc tcttattgta
     1141 acagctttaa gggccaattc tgctgtcaaa ttagaaaacc tttactttca gtcgatggat
     1201 tataaggacc acgatggaga ctacaaagat catgacattg attacaagga cgacgatgac
     1261 aaataagcta gcttgactga ctgagataca gcgtaccttc agctcacaga catgataaga
     1321 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
     1381 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttaac
     1441 aacaacaatt gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa
     1501 agcaagtaaa acctctacaa atgtggtatt ggcccatctc tatcggtatc gtagcataac
     1561 cccttggggc ctctaaacgg gtcttgaggg gttttttgtg cccctcgggc cggattgcta
     1621 tctaccggca ttggcgcaga aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca
     1681 catatgccag attcagcaac ggatacggct tccccaactt gcccacttcc atacgtgtcc
     1741 tccttaccag aaatttatcc ttaaggtcgt cagctatcct gcaggcgatc tctcgatttc
     1801 gatcaagaca ttcctttaat ggtcttttct ggacaccact aggggtcaga agtagttcat
     1861 caaactttct tccctcccta atctcattgg ttaccttggg ctatcgaaac ttaattaacc
     1921 agtcaagtca gctacttggc gagatcgact tgtctgggtt tcgactacgc tcagaattgc
     1981 gtcagtcaag ttcgatctgg tccttgctat tgcacccgtt ctccgattac gagtttcatt
     2041 taaatcatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     2101 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     2161 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     2221 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     2281 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     2341 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     2401 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     2461 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     2521 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     2581 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     2641 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     2701 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     2761 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     2821 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     2881 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc atttaaattt
     2941 ccgaactctc caaggccctc gtcggaaaat cttcaaacct ttcgtccgat ccatcttgca
     3001 ggctacctct cgaacgaact atcgcaagtc tcttggccgg ccttgcgcct tggctattgc
     3061 ttggcagcgc ctatcgccag gtattactcc aatcccgaat atccgagatc gggatcaccc
     3121 gagagaagtt caacctacat cctcaatccc gatctatccg agatccgagg aatatcgaaa
     3181 tcggggcgcg cctggcctcc gcgccgggtt ttggcgcctc ccgcgggcgc ccccctcgtc
     3241 acggcgagcg ctgccacgtc agacgaaggg cgcaggagcg tcctgatcct tccgcccgga
     3301 cgctcaggac agcggcccgc tgctcataag actcggcctt agaaccccag tatcagcaga
     3361 aggacatttt aggacgggac ttgggtgact ctagggcact ggttttcttt ccagagagcg
     3421 gaacaggcga ggaaaagtag tcccttctcg gcgattctgc ggagggatct ccgtggggcg
     3481 gtgaacgccg atgattatat aaggacgcgc cgggtgtggc acagctagtt ccgtcgcagc
     3541 cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg gtgagtagcg
     3601 ggctgctggg ctggccgggg ctttcgtggc cgccgggccg ctcggtggga cggaagcgtg
     3661 tggagagacc gccaagggct gtagtctggg tccgcgagca aggttgccct gaactggggg
     3721 ttggggggag cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag acgcttgtga
     3781 ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa gaacccaagg
     3841 tcttgagccc ttcgctaatg cgggaaagct cttattcggg tgagatgggc tgggcaccat
     3901 ctggggaccc tgacgtgaag tttgtcactg actggagaac tcggtttgtc gtctgttgcg
     3961 ggggcggcag ttatggcggt gccgttgggc agtgcacccg tacctttggg agcgcgcgcc
     4021 ctcgtcgtgt cgtgacgtca cccgttctgt tggcttataa tgcagggtgg ggccacctgc
     4081 cggtaggtgt gcggtaggct tttctccgtc gcaggacgca gggttcgggc ctagggtagg
     4141 ctctcctgaa tcgacaggcg ccggacctct ggtgagggga gggataagtg aggcgtcagt
     4201 ttctttggtc ggttttatgt acctatcttc ttaagtagct gaagctccgg ttttgaacta
     4261 tgcgctcggg gttggcgagt gtgttttgtg aagtttttta ggcacctttt gaaatgtaat
     4321 catttgggtc aatatgtaat tttcagtgtt agacttgtaa attgtccgct aaattctggc
     4381 cgtttttggc ttttttgtta gacaacatga ccgagtacaa gcccacggtg cgcctcgcca
     4441 cccgcgacga cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg
     4501 ccacgcgcca caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac
     4561 tcttcctcac gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg
     4621 cggtggcggt ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg
     4681 gcccgcgcat ggccgagttg agcggttccc ggctggccgc gcagcaacag atggagggcc
     4741 tcctggcgcc gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc
     4801 ccgaccacca gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggccgccg
     4861 agcgcgccgg ggtgcccgcc ttcctggaga cctccgcgcc ccgcaacctc cccttctacg
     4921 agcggctcgg cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt
     4981 gcatgacccg caagcccggt gcctaggcta gtatgtaagc ctagtcttag ataataaaat
     5041 cgctatccat cgaagatgga tgtgtgttgg ttttttgtgt gtgtaacgct aggcgcgcct
     5101 ggtgtaccga gaacgatcct ctcagtgcga gtctcgacga tccatatcgt tgcttggcag
     5161 tcagccagtc ggaatccagc ttgggaccca ggaagtccaa tcgtcagata ttgtactcaa
     5221 gcctggtcac ggcagcgtac cgatctgttt aaacctagat attgatagtc tgatcggtca
     5281 acgtataatc gagtcctagc ttttgcaaac atctatcaag agacaggatc agcaggaggc
     5341 tttcgcatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     5401 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5461 ggtgcgcgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5521 cgccccgaag aacgctttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5581 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5641 gacttggttg agtattcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5701 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5761 acgattggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5821 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5881 acgatgcctg tagcaatggc aacaaccttg cgtaaactat taactggcga actacttact
     5941 ctagcttccc ggcaacagtt gatagactgg atggaggcgg ataaagttgc aggaccactt
     6001 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6061 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6121 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6181 ggtgcctcac tgattaagca ttggtaaccg attctaggtg cattggcgca gaaaaaaatg
     6241 cctgatgcga cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg
     6301 cttccccaac ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaagatc
     6361 ccgaatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga gcttacgcga
     6421 acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc gtcagcttac
     6481 ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca ggtatcttct
     6541 tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa cccctcaaga
     6601 cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac acaaaaaacc
     6661 aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat cgagtgtagc
     6721 cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccg
//
