LOCUS       sequence-295713-        7196 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 7196)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7196)
  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-m4133164
FEATURES             Location/Qualifiers
     source          1..7196
                     /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..667
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    736..857
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(747..766)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     801..820
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      972..1018
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1048..1184
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1229..1300
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1526..2114)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1606..1625)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2614..3823
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3660..3679
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3827..4855
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    4882..4930
                     /note="synthetic polyadenylation signal"
     CDS             join(5196..5264,5265..6056)
                     /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(5414..5433)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6069..6205
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6350..6421
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6426..6472)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6490..6538
                     /note="synthetic polyadenylation signal"
     enhancer        6619..6922
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6924..7126
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7077..7097
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7195..7196,1..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 cgaaagaaac cgcggcagct aagttcgaga ggcagcacat
      661 ggacagttaa ggatccgcag gcctctgcta gcttgactga ctgagataca gcgtaccttc
      721 agctcacaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
      781 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
      841 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
      901 aggtgtggga ggttttttaa agcaagtaaa acctctacaa atgtggtatt ggcccatctc
      961 tatcggtatc gtagcataac cccttggggc ctctaaacgg gtcttgaggg gttttttgtg
     1021 cccctcgggc cggattgcta tctaccggca ttggcgcaga aaaaaatgcc tgatgcgacg
     1081 ctgcgcgtct tatactccca catatgccag attcagcaac ggatacggct tccccaactt
     1141 gcccacttcc atacgtgtcc tccttaccag aaatttatcc ttaaggtcgt cagctatcct
     1201 gcaggcgatc tctcgatttc gatcaagaca ttcctttaat ggtcttttct ggacaccact
     1261 aggggtcaga agtagttcat caaactttct tccctcccta atctcattgg ttaccttggg
     1321 ctatcgaaac ttaattaacc agtcaagtca gctacttggc gagatcgact tgtctgggtt
     1381 tcgactacgc tcagaattgc gtcagtcaag ttcgatctgg tccttgctat tgcacccgtt
     1441 ctccgattac gagtttcatt taaatcatgt gagcaaaagg ccagcaaaag gccaggaacc
     1501 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     1561 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     1621 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     1681 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     1741 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     1801 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     1861 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     1921 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     1981 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     2041 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     2101 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     2161 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     2221 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     2281 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     2341 ccatagttgc atttaaattt ccgaactctc caaggccctc gtcggaaaat cttcaaacct
     2401 ttcgtccgat ccatcttgca ggctacctct cgaacgaact atcgcaagtc tcttggccgg
     2461 ccttgcgcct tggctattgc ttggcagcgc ctatcgccag gtattactcc aatcccgaat
     2521 atccgagatc gggatcaccc gagagaagtt caacctacat cctcaatccc gatctatccg
     2581 agatccgagg aatatcgaaa tcggggcgcg cctggcctcc gcgccgggtt ttggcgcctc
     2641 ccgcgggcgc ccccctcgtc acggcgagcg ctgccacgtc agacgaaggg cgcaggagcg
     2701 tcctgatcct tccgcccgga cgctcaggac agcggcccgc tgctcataag actcggcctt
     2761 agaaccccag tatcagcaga aggacatttt aggacgggac ttgggtgact ctagggcact
     2821 ggttttcttt ccagagagcg gaacaggcga ggaaaagtag tcccttctcg gcgattctgc
     2881 ggagggatct ccgtggggcg gtgaacgccg atgattatat aaggacgcgc cgggtgtggc
     2941 acagctagtt ccgtcgcagc cgggatttgg gtcgcggttc ttgtttgtgg atcgctgtga
     3001 tcgtcacttg gtgagtagcg ggctgctggg ctggccgggg ctttcgtggc cgccgggccg
     3061 ctcggtggga cggaagcgtg tggagagacc gccaagggct gtagtctggg tccgcgagca
     3121 aggttgccct gaactggggg ttggggggag cgcagcaaaa tggcggctgt tcccgagtct
     3181 tgaatggaag acgcttgtga ggcgggctgt gaggtcgttg aaacaaggtg gggggcatgg
     3241 tgggcggcaa gaacccaagg tcttgagccc ttcgctaatg cgggaaagct cttattcggg
     3301 tgagatgggc tgggcaccat ctggggaccc tgacgtgaag tttgtcactg actggagaac
     3361 tcggtttgtc gtctgttgcg ggggcggcag ttatggcggt gccgttgggc agtgcacccg
     3421 tacctttggg agcgcgcgcc ctcgtcgtgt cgtgacgtca cccgttctgt tggcttataa
     3481 tgcagggtgg ggccacctgc cggtaggtgt gcggtaggct tttctccgtc gcaggacgca
     3541 gggttcgggc ctagggtagg ctctcctgaa tcgacaggcg ccggacctct ggtgagggga
     3601 gggataagtg aggcgtcagt ttctttggtc ggttttatgt acctatcttc ttaagtagct
     3661 gaagctccgg ttttgaacta tgcgctcggg gttggcgagt gtgttttgtg aagtttttta
     3721 ggcacctttt gaaatgtaat catttgggtc aatatgtaat tttcagtgtt agacttgtaa
     3781 attgtccgct aaattctggc cgtttttggc ttttttgtta gacaacatgg gtaaaaagcc
     3841 tgaactcacc gcgacgtctg tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga
     3901 cctgatgcag ctctcggagg gcgaagaatc tcgtgctttc agcttcgatg taggagggcg
     3961 tggatatgtc ctgcgggtaa atagctgcgc cgatggtttc tacaaagatc gttatgttta
     4021 tcggcacttt gcatccgccg cgctcccgat tccggaagtg cttgacattg gggagttcag
     4081 cgagagcctg acctattgca tctcccgccg tgcacagggt gtcacgttgc aagacctgcc
     4141 tgaaaccgaa ctgcccgctg ttctgcagcc ggtcgcggag gcaatggatg ccatcgctgc
     4201 cgccgatctt agccagacga gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata
     4261 cactacatgg cgtgatttca tatgcgcgat tgctgatccc catgtgtatc actggcaaac
     4321 tgtgatggac gacaccgtca gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg
     4381 ggccgaggac tgccccgaag tccggcacct cgtgcacgcg gatttcggct ccaacaatgt
     4441 cctgacggac aatggccgca taacagcggt cattgactgg agcgaggcga tgttcgggga
     4501 ttcccaatac gaggtcgcca acatcttctt ctggaggccg tggttggctt gtatggagca
     4561 gcagacgcgc tacttcgagc ggaggcatcc ggagcttgca ggatcgcccc ggctccgggc
     4621 gtatatgctc cgcattggtc ttgaccaact ctatcagagc ttggttgacg gcaatttcga
     4681 tgatgcagct tgggcgcagg gtcgatgcga cgcaatcgtc cgatccggag ccgggactgt
     4741 cgggcgtaca caaatcgccc gcagaagcgc cgccgtctgg accgatggct gtgtagaagt
     4801 actcgccgat agtggaaacc gacgccccag cactcgtccg agggcaaagg aataagctag
     4861 tatgtaagcc tagtcttaga taataaaatc gctatccatc gaagatggat gtgtgttggt
     4921 tttttgtgtg tgtaacgcta ggcgcgcctg gtgtaccgag aacgatcctc tcagtgcgag
     4981 tctcgacgat ccatatcgtt gcttggcagt cagccagtcg gaatccagct tgggacccag
     5041 gaagtccaat cgtcagatat tgtactcaag cctggtcacg gcagcgtacc gatctgttta
     5101 aacctagata ttgatagtct gatcggtcaa cgtataatcg agtcctagct tttgcaaaca
     5161 tctatcaaga gacaggatca gcaggaggct ttcgcatgag tattcaacat ttccgtgtcg
     5221 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
     5281 tgaaagtaaa agatgctgaa gatcagttgg gtgcgcgagt gggttacatc gaactggatc
     5341 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgctttcca atgatgagca
     5401 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
     5461 tcggtcgccg catacactat tctcagaatg acttggttga gtattcacca gtcacagaaa
     5521 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
     5581 ataacactgc ggccaactta cttctgacaa cgattggagg accgaaggag ctaaccgctt
     5641 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
     5701 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaaccttgc
     5761 gtaaactatt aactggcgaa ctacttactc tagcttcccg gcaacagttg atagactgga
     5821 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
     5881 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
     5941 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
     6001 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaaccga
     6061 ttctaggtgc attggcgcag aaaaaaatgc ctgatgcgac gctgcgcgtc ttatactccc
     6121 acatatgcca gattcagcaa cggatacggc ttccccaact tgcccacttc catacgtgtc
     6181 ctccttacca gaaatttatc cttaagatcg tttaaactcg actctggctc tatcgaatct
     6241 ccgtcgtttc gagcttacgc gaacagccgt ggcgctcatt tgctcgtcgg gcatcgaatc
     6301 tcgtcagcta tcgtcagctt acctttttgg cagcgatcgc ggctcccgac atcttggacc
     6361 attagctcca caggtatctt cttccctcta gtggtcataa cagcagcttc agctacctct
     6421 caattcaaaa aacccctcaa gacccgttta gaggccccaa ggggttatgc tatcaatcgt
     6481 tgcgttacac acacaaaaaa ccaacacaca tccatcttcg atggatagcg attttattat
     6541 ctaactgctg atcgagtgta gccagatcta gtaatcaatt acggggtcat tagttcatag
     6601 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     6661 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     6721 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     6781 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     6841 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     6901 attagtcatc gctattacca tgctgatgcg gttttggcag tacatcaatg ggcgtggata
     6961 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     7021 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
     7081 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg
     7141 tcagatcaga tctttgtcga tcctaccatc cactcgacac acccgccagc ggccgc
//
