LOCUS       sequence-295383-        7706 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 7706)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7706)
  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-m4134227
FEATURES             Location/Qualifiers
     source          1..7706
                     /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             656..733
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     CDS             761..781
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    1246..1367
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1257..1276)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1311..1330
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1482..1528
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1558..1694
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1739..1810
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2036..2624)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2116..2135)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3124..4333
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4170..4189
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4337..5365
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5392..5440
                     /note="synthetic polyadenylation signal"
     CDS             join(5706..5774,5775..6566)
                     /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(5924..5943)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6579..6715
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6860..6931
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6936..6982)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7000..7048
                     /note="synthetic polyadenylation signal"
     enhancer        7129..7432
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7434..7636
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7587..7607
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7705..7706,1..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 cgggacggcg aatcccgggg ctgataaatc cctggaaaag
      661 gagatggaag aagaacttta ttgcagtgtc tgcggctaac cggttcaaaa agatcagttc
      721 aagcggcgcc ctagattacg acattcctac gaccgcctcc gagaatctct atttccaggg
      781 tgaatttggc ttggcacaac atgacgaggc cgtcgataac aagtttaata aagaacagca
      841 gaacgctttc tatgagatcc ttcacttacc aaatctgaac gaagagcaac gcaacgcttt
      901 cattcagagc ctgaaagacg atccctcgca gtctgcaaat ctcctggccg aagccaagaa
      961 gctgaatgac gcacaggctc caaaagttga taacaagttt aacaaggagc agcagaatgc
     1021 tttctacgag atattgcacc ttcctaatct gaatgaggaa caaagaaatg catttatcca
     1081 gtccctaaaa gacgatccat cacagagtgc caacctgctt gcggaagcca aaaagctcaa
     1141 tgatgcccag gccccgaaag tagacgcgaa ccatcaataa ggatccgcag gcctctgcta
     1201 gcttgactga ctgagataca gcgtaccttc agctcacaga catgataaga tacattgatg
     1261 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     1321 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     1381 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa
     1441 acctctacaa atgtggtatt ggcccatctc tatcggtatc gtagcataac cccttggggc
     1501 ctctaaacgg gtcttgaggg gttttttgtg cccctcgggc cggattgcta tctaccggca
     1561 ttggcgcaga aaaaaatgcc tgatgcgacg ctgcgcgtct tatactccca catatgccag
     1621 attcagcaac ggatacggct tccccaactt gcccacttcc atacgtgtcc tccttaccag
     1681 aaatttatcc ttaaggtcgt cagctatcct gcaggcgatc tctcgatttc gatcaagaca
     1741 ttcctttaat ggtcttttct ggacaccact aggggtcaga agtagttcat caaactttct
     1801 tccctcccta atctcattgg ttaccttggg ctatcgaaac ttaattaacc agtcaagtca
     1861 gctacttggc gagatcgact tgtctgggtt tcgactacgc tcagaattgc gtcagtcaag
     1921 ttcgatctgg tccttgctat tgcacccgtt ctccgattac gagtttcatt taaatcatgt
     1981 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     2041 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     2101 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     2161 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     2221 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     2281 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     2341 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     2401 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     2461 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     2521 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     2581 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     2641 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     2701 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     2761 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     2821 ctatctcagc gatctgtcta tttcgttcat ccatagttgc atttaaattt ccgaactctc
     2881 caaggccctc gtcggaaaat cttcaaacct ttcgtccgat ccatcttgca ggctacctct
     2941 cgaacgaact atcgcaagtc tcttggccgg ccttgcgcct tggctattgc ttggcagcgc
     3001 ctatcgccag gtattactcc aatcccgaat atccgagatc gggatcaccc gagagaagtt
     3061 caacctacat cctcaatccc gatctatccg agatccgagg aatatcgaaa tcggggcgcg
     3121 cctggcctcc gcgccgggtt ttggcgcctc ccgcgggcgc ccccctcgtc acggcgagcg
     3181 ctgccacgtc agacgaaggg cgcaggagcg tcctgatcct tccgcccgga cgctcaggac
     3241 agcggcccgc tgctcataag actcggcctt agaaccccag tatcagcaga aggacatttt
     3301 aggacgggac ttgggtgact ctagggcact ggttttcttt ccagagagcg gaacaggcga
     3361 ggaaaagtag tcccttctcg gcgattctgc ggagggatct ccgtggggcg gtgaacgccg
     3421 atgattatat aaggacgcgc cgggtgtggc acagctagtt ccgtcgcagc cgggatttgg
     3481 gtcgcggttc ttgtttgtgg atcgctgtga tcgtcacttg gtgagtagcg ggctgctggg
     3541 ctggccgggg ctttcgtggc cgccgggccg ctcggtggga cggaagcgtg tggagagacc
     3601 gccaagggct gtagtctggg tccgcgagca aggttgccct gaactggggg ttggggggag
     3661 cgcagcaaaa tggcggctgt tcccgagtct tgaatggaag acgcttgtga ggcgggctgt
     3721 gaggtcgttg aaacaaggtg gggggcatgg tgggcggcaa gaacccaagg tcttgagccc
     3781 ttcgctaatg cgggaaagct cttattcggg tgagatgggc tgggcaccat ctggggaccc
     3841 tgacgtgaag tttgtcactg actggagaac tcggtttgtc gtctgttgcg ggggcggcag
     3901 ttatggcggt gccgttgggc agtgcacccg tacctttggg agcgcgcgcc ctcgtcgtgt
     3961 cgtgacgtca cccgttctgt tggcttataa tgcagggtgg ggccacctgc cggtaggtgt
     4021 gcggtaggct tttctccgtc gcaggacgca gggttcgggc ctagggtagg ctctcctgaa
     4081 tcgacaggcg ccggacctct ggtgagggga gggataagtg aggcgtcagt ttctttggtc
     4141 ggttttatgt acctatcttc ttaagtagct gaagctccgg ttttgaacta tgcgctcggg
     4201 gttggcgagt gtgttttgtg aagtttttta ggcacctttt gaaatgtaat catttgggtc
     4261 aatatgtaat tttcagtgtt agacttgtaa attgtccgct aaattctggc cgtttttggc
     4321 ttttttgtta gacaacatgg gtaaaaagcc tgaactcacc gcgacgtctg tcgagaagtt
     4381 tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg gcgaagaatc
     4441 tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa atagctgcgc
     4501 cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatccgccg cgctcccgat
     4561 tccggaagtg cttgacattg gggagttcag cgagagcctg acctattgca tctcccgccg
     4621 tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg ttctgcagcc
     4681 ggtcgcggag gcaatggatg ccatcgctgc cgccgatctt agccagacga gcgggttcgg
     4741 cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca tatgcgcgat
     4801 tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca gtgcgtccgt
     4861 cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag tccggcacct
     4921 cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca taacagcggt
     4981 cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca acatcttctt
     5041 ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc ggaggcatcc
     5101 ggagcttgca ggatcgcccc ggctccgggc gtatatgctc cgcattggtc ttgaccaact
     5161 ctatcagagc ttggttgacg gcaatttcga tgatgcagct tgggcgcagg gtcgatgcga
     5221 cgcaatcgtc cgatccggag ccgggactgt cgggcgtaca caaatcgccc gcagaagcgc
     5281 cgccgtctgg accgatggct gtgtagaagt actcgccgat agtggaaacc gacgccccag
     5341 cactcgtccg agggcaaagg aataagctag tatgtaagcc tagtcttaga taataaaatc
     5401 gctatccatc gaagatggat gtgtgttggt tttttgtgtg tgtaacgcta ggcgcgcctg
     5461 gtgtaccgag aacgatcctc tcagtgcgag tctcgacgat ccatatcgtt gcttggcagt
     5521 cagccagtcg gaatccagct tgggacccag gaagtccaat cgtcagatat tgtactcaag
     5581 cctggtcacg gcagcgtacc gatctgttta aacctagata ttgatagtct gatcggtcaa
     5641 cgtataatcg agtcctagct tttgcaaaca tctatcaaga gacaggatca gcaggaggct
     5701 ttcgcatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5761 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5821 gtgcgcgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5881 gccccgaaga acgctttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5941 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     6001 acttggttga gtattcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     6061 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     6121 cgattggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     6181 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     6241 cgatgcctgt agcaatggca acaaccttgc gtaaactatt aactggcgaa ctacttactc
     6301 tagcttcccg gcaacagttg atagactgga tggaggcgga taaagttgca ggaccacttc
     6361 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     6421 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6481 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6541 gtgcctcact gattaagcat tggtaaccga ttctaggtgc attggcgcag aaaaaaatgc
     6601 ctgatgcgac gctgcgcgtc ttatactccc acatatgcca gattcagcaa cggatacggc
     6661 ttccccaact tgcccacttc catacgtgtc ctccttacca gaaatttatc cttaagatcg
     6721 tttaaactcg actctggctc tatcgaatct ccgtcgtttc gagcttacgc gaacagccgt
     6781 ggcgctcatt tgctcgtcgg gcatcgaatc tcgtcagcta tcgtcagctt acctttttgg
     6841 cagcgatcgc ggctcccgac atcttggacc attagctcca caggtatctt cttccctcta
     6901 gtggtcataa cagcagcttc agctacctct caattcaaaa aacccctcaa gacccgttta
     6961 gaggccccaa ggggttatgc tatcaatcgt tgcgttacac acacaaaaaa ccaacacaca
     7021 tccatcttcg atggatagcg attttattat ctaactgctg atcgagtgta gccagatcta
     7081 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
     7141 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
     7201 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
     7261 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
     7321 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
     7381 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tgctgatgcg
     7441 gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct
     7501 ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa
     7561 atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt
     7621 ctatataagc agagctggtt tagtgaaccg tcagatcaga tctttgtcga tcctaccatc
     7681 cactcgacac acccgccagc ggccgc
//
