LOCUS       sequence-295764-        7190 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 7190)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7190)
  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-m4133929
FEATURES             Location/Qualifiers
     source          1..7190
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             596..616
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             617..661
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    730..851
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(741..760)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     795..814
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      966..1012
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1042..1178
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1223..1294
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1520..2108)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1600..1619)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2608..3817
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3654..3673
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3821..4849
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    4876..4924
                     /note="synthetic polyadenylation signal"
     CDS             join(5190..5258,5259..6050)
                     /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(5408..5427)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6063..6199
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6344..6415
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6420..6466)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6484..6532
                     /note="synthetic polyadenylation signal"
     enhancer        6613..6916
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6918..7120
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7071..7091
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgata gcctcagagt ttagttccct tccatcatat gcagcttttg ctactgctca
       61 agaagcttat gagcaggctg ttgctaatgg tgattctgaa gttgttctta aaaagttgaa
      121 gaagtctttg aatgtggcta aatctgaatt tgaccgtgat gcagccatgc aacgtaagtt
      181 ggaaaagatg gctgatcaag ctatgaccca aatgtataaa caggctagat ctgaggacaa
      241 gagggcaaaa gttactagtg ctatgcagac aatgcttttc actatgctta gaaagttgga
      301 taatgatgca ctcaacaaca ttatcaacaa tgcaagagat ggttgtgttc ccttgaacat
      361 aatacctctt acaacagcag ccaaactaat ggttgtcata ccagactata acacatataa
      421 aaatacgtgt gatggtacaa catttactta tgcatcagca ttgtgggaaa tccaacaggt
      481 tgtagatgca gatagtaaaa ttgttcaact tagtgaaatt agtatggaca attcacctaa
      541 tttagcatgg cctcttattg taacagcttt aagggccaat tctgctgtca aattagaaaa
      601 cctttacttt cagtcgaaag aaaccgcggc agctaagttc gagaggcagc acatggacag
      661 ttaaggatcc gcaggcctct gctagcttga ctgactgaga tacagcgtac cttcagctca
      721 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
      781 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
      841 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggaggtgt
      901 gggaggtttt ttaaagcaag taaaacctct acaaatgtgg tattggccca tctctatcgg
      961 tatcgtagca taaccccttg gggcctctaa acgggtcttg aggggttttt tgtgcccctc
     1021 gggccggatt gctatctacc ggcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc
     1081 gtcttatact cccacatatg ccagattcag caacggatac ggcttcccca acttgcccac
     1141 ttccatacgt gtcctcctta ccagaaattt atccttaagg tcgtcagcta tcctgcaggc
     1201 gatctctcga tttcgatcaa gacattcctt taatggtctt ttctggacac cactaggggt
     1261 cagaagtagt tcatcaaact ttcttccctc cctaatctca ttggttacct tgggctatcg
     1321 aaacttaatt aaccagtcaa gtcagctact tggcgagatc gacttgtctg ggtttcgact
     1381 acgctcagaa ttgcgtcagt caagttcgat ctggtccttg ctattgcacc cgttctccga
     1441 ttacgagttt catttaaatc atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     1501 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     1561 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     1621 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     1681 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     1741 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     1801 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     1861 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     1921 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     1981 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     2041 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     2101 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     2161 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     2221 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     2281 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     2341 ttgcatttaa atttccgaac tctccaaggc cctcgtcgga aaatcttcaa acctttcgtc
     2401 cgatccatct tgcaggctac ctctcgaacg aactatcgca agtctcttgg ccggccttgc
     2461 gccttggcta ttgcttggca gcgcctatcg ccaggtatta ctccaatccc gaatatccga
     2521 gatcgggatc acccgagaga agttcaacct acatcctcaa tcccgatcta tccgagatcc
     2581 gaggaatatc gaaatcgggg cgcgcctggc ctccgcgccg ggttttggcg cctcccgcgg
     2641 gcgcccccct cgtcacggcg agcgctgcca cgtcagacga agggcgcagg agcgtcctga
     2701 tccttccgcc cggacgctca ggacagcggc ccgctgctca taagactcgg ccttagaacc
     2761 ccagtatcag cagaaggaca ttttaggacg ggacttgggt gactctaggg cactggtttt
     2821 ctttccagag agcggaacag gcgaggaaaa gtagtccctt ctcggcgatt ctgcggaggg
     2881 atctccgtgg ggcggtgaac gccgatgatt atataaggac gcgccgggtg tggcacagct
     2941 agttccgtcg cagccgggat ttgggtcgcg gttcttgttt gtggatcgct gtgatcgtca
     3001 cttggtgagt agcgggctgc tgggctggcc ggggctttcg tggccgccgg gccgctcggt
     3061 gggacggaag cgtgtggaga gaccgccaag ggctgtagtc tgggtccgcg agcaaggttg
     3121 ccctgaactg ggggttgggg ggagcgcagc aaaatggcgg ctgttcccga gtcttgaatg
     3181 gaagacgctt gtgaggcggg ctgtgaggtc gttgaaacaa ggtggggggc atggtgggcg
     3241 gcaagaaccc aaggtcttga gcccttcgct aatgcgggaa agctcttatt cgggtgagat
     3301 gggctgggca ccatctgggg accctgacgt gaagtttgtc actgactgga gaactcggtt
     3361 tgtcgtctgt tgcgggggcg gcagttatgg cggtgccgtt gggcagtgca cccgtacctt
     3421 tgggagcgcg cgccctcgtc gtgtcgtgac gtcacccgtt ctgttggctt ataatgcagg
     3481 gtggggccac ctgccggtag gtgtgcggta ggcttttctc cgtcgcagga cgcagggttc
     3541 gggcctaggg taggctctcc tgaatcgaca ggcgccggac ctctggtgag gggagggata
     3601 agtgaggcgt cagtttcttt ggtcggtttt atgtacctat cttcttaagt agctgaagct
     3661 ccggttttga actatgcgct cggggttggc gagtgtgttt tgtgaagttt tttaggcacc
     3721 ttttgaaatg taatcatttg ggtcaatatg taattttcag tgttagactt gtaaattgtc
     3781 cgctaaattc tggccgtttt tggctttttt gttagacaac atgggtaaaa agcctgaact
     3841 caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct ccgacctgat
     3901 gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag ggcgtggata
     3961 tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg tttatcggca
     4021 ctttgcatcc gccgcgctcc cgattccgga agtgcttgac attggggagt tcagcgagag
     4081 cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc tgcctgaaac
     4141 cgaactgccc gctgttctgc agccggtcgc ggaggcaatg gatgccatcg ctgccgccga
     4201 tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc aatacactac
     4261 atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc aaactgtgat
     4321 ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc tttgggccga
     4381 ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca atgtcctgac
     4441 ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg gggattccca
     4501 atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg agcagcagac
     4561 gcgctacttc gagcggaggc atccggagct tgcaggatcg ccccggctcc gggcgtatat
     4621 gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt tcgatgatgc
     4681 agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga ctgtcgggcg
     4741 tacacaaatc gcccgcagaa gcgccgccgt ctggaccgat ggctgtgtag aagtactcgc
     4801 cgatagtgga aaccgacgcc ccagcactcg tccgagggca aaggaataag ctagtatgta
     4861 agcctagtct tagataataa aatcgctatc catcgaagat ggatgtgtgt tggttttttg
     4921 tgtgtgtaac gctaggcgcg cctggtgtac cgagaacgat cctctcagtg cgagtctcga
     4981 cgatccatat cgttgcttgg cagtcagcca gtcggaatcc agcttgggac ccaggaagtc
     5041 caatcgtcag atattgtact caagcctggt cacggcagcg taccgatctg tttaaaccta
     5101 gatattgata gtctgatcgg tcaacgtata atcgagtcct agcttttgca aacatctatc
     5161 aagagacagg atcagcagga ggctttcgca tgagtattca acatttccgt gtcgccctta
     5221 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5281 taaaagatgc tgaagatcag ttgggtgcgc gagtgggtta catcgaactg gatctcaaca
     5341 gcggtaagat ccttgagagt tttcgccccg aagaacgctt tccaatgatg agcactttta
     5401 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     5461 gccgcataca ctattctcag aatgacttgg ttgagtattc accagtcaca gaaaagcatc
     5521 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     5581 ctgcggccaa cttacttctg acaacgattg gaggaccgaa ggagctaacc gcttttttgc
     5641 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     5701 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacc ttgcgtaaac
     5761 tattaactgg cgaactactt actctagctt cccggcaaca gttgatagac tggatggagg
     5821 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     5881 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     5941 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     6001 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ccgattctag
     6061 gtgcattggc gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac tcccacatat
     6121 gccagattca gcaacggata cggcttcccc aacttgccca cttccatacg tgtcctcctt
     6181 accagaaatt tatccttaag atcgtttaaa ctcgactctg gctctatcga atctccgtcg
     6241 tttcgagctt acgcgaacag ccgtggcgct catttgctcg tcgggcatcg aatctcgtca
     6301 gctatcgtca gcttaccttt ttggcagcga tcgcggctcc cgacatcttg gaccattagc
     6361 tccacaggta tcttcttccc tctagtggtc ataacagcag cttcagctac ctctcaattc
     6421 aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctatcaa tcgttgcgtt
     6481 acacacacaa aaaaccaaca cacatccatc ttcgatggat agcgatttta ttatctaact
     6541 gctgatcgag tgtagccaga tctagtaatc aattacgggg tcattagttc atagcccata
     6601 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
     6661 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
     6721 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     6781 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     6841 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     6901 catcgctatt accatgctga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
     6961 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
     7021 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
     7081 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
     7141 cagatctttg tcgatcctac catccactcg acacacccgc cagcggccgc
//
