LOCUS       sequence-295909-        7541 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 7541)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7541)
  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-m4134471
FEATURES             Location/Qualifiers
     source          1..7541
                     /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             971..988
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    1081..1202
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1092..1111)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1146..1165
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1317..1363
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1393..1529
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1574..1645
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1871..2459)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1951..1970)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2959..4168
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4005..4024
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4172..5200
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5227..5275
                     /note="synthetic polyadenylation signal"
     CDS             join(5541..5609,5610..6401)
                     /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(5759..5778)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6414..6550
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6695..6766
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6771..6817)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6835..6883
                     /note="synthetic polyadenylation signal"
     enhancer        6964..7267
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7269..7471
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7422..7442
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      join(7540..7541,1..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 cgatgagtga caaaatcatt catctaaccg atgactcctt
      661 tgatacagac gtgttgaagg ccgatggtgc gatactggta gatttctggg ctgaatggtg
      721 tggcccgtgc aagatgatcg cacccattct ggacgagatc gccgacgagt atcaaggaaa
      781 attaactgtc gctaagctta atatagatca gaacccaggg acggcaccta aatacggcat
      841 caggggaata cctaccctcc tgctctttaa gaatggggaa gttgccgcga caaaagtggg
      901 agccctgtct aagggccagc taaaagagtt cttggacgct aacctggcag gttcaggctc
      961 ggggcacatg caccatcacc atcaccatag cagcgggctc gtgccacgcg gctaaggatc
     1021 cgcaggcctc tgctagcttg actgactgag atacagcgta ccttcagctc acagacatga
     1081 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     1141 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     1201 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg tgggaggttt
     1261 tttaaagcaa gtaaaacctc tacaaatgtg gtattggccc atctctatcg gtatcgtagc
     1321 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgtgcccct cgggccggat
     1381 tgctatctac cggcattggc gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac
     1441 tcccacatat gccagattca gcaacggata cggcttcccc aacttgccca cttccatacg
     1501 tgtcctcctt accagaaatt tatccttaag gtcgtcagct atcctgcagg cgatctctcg
     1561 atttcgatca agacattcct ttaatggtct tttctggaca ccactagggg tcagaagtag
     1621 ttcatcaaac tttcttccct ccctaatctc attggttacc ttgggctatc gaaacttaat
     1681 taaccagtca agtcagctac ttggcgagat cgacttgtct gggtttcgac tacgctcaga
     1741 attgcgtcag tcaagttcga tctggtcctt gctattgcac ccgttctccg attacgagtt
     1801 tcatttaaat catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     1861 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     1921 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     1981 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     2041 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     2101 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     2161 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     2221 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     2281 agtggtggcc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     2341 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     2401 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     2461 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     2521 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     2581 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     2641 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcattta
     2701 aatttccgaa ctctccaagg ccctcgtcgg aaaatcttca aacctttcgt ccgatccatc
     2761 ttgcaggcta cctctcgaac gaactatcgc aagtctcttg gccggccttg cgccttggct
     2821 attgcttggc agcgcctatc gccaggtatt actccaatcc cgaatatccg agatcgggat
     2881 cacccgagag aagttcaacc tacatcctca atcccgatct atccgagatc cgaggaatat
     2941 cgaaatcggg gcgcgcctgg cctccgcgcc gggttttggc gcctcccgcg ggcgcccccc
     3001 tcgtcacggc gagcgctgcc acgtcagacg aagggcgcag gagcgtcctg atccttccgc
     3061 ccggacgctc aggacagcgg cccgctgctc ataagactcg gccttagaac cccagtatca
     3121 gcagaaggac attttaggac gggacttggg tgactctagg gcactggttt tctttccaga
     3181 gagcggaaca ggcgaggaaa agtagtccct tctcggcgat tctgcggagg gatctccgtg
     3241 gggcggtgaa cgccgatgat tatataagga cgcgccgggt gtggcacagc tagttccgtc
     3301 gcagccggga tttgggtcgc ggttcttgtt tgtggatcgc tgtgatcgtc acttggtgag
     3361 tagcgggctg ctgggctggc cggggctttc gtggccgccg ggccgctcgg tgggacggaa
     3421 gcgtgtggag agaccgccaa gggctgtagt ctgggtccgc gagcaaggtt gccctgaact
     3481 gggggttggg gggagcgcag caaaatggcg gctgttcccg agtcttgaat ggaagacgct
     3541 tgtgaggcgg gctgtgaggt cgttgaaaca aggtgggggg catggtgggc ggcaagaacc
     3601 caaggtcttg agcccttcgc taatgcggga aagctcttat tcgggtgaga tgggctgggc
     3661 accatctggg gaccctgacg tgaagtttgt cactgactgg agaactcggt ttgtcgtctg
     3721 ttgcgggggc ggcagttatg gcggtgccgt tgggcagtgc acccgtacct ttgggagcgc
     3781 gcgccctcgt cgtgtcgtga cgtcacccgt tctgttggct tataatgcag ggtggggcca
     3841 cctgccggta ggtgtgcggt aggcttttct ccgtcgcagg acgcagggtt cgggcctagg
     3901 gtaggctctc ctgaatcgac aggcgccgga cctctggtga ggggagggat aagtgaggcg
     3961 tcagtttctt tggtcggttt tatgtaccta tcttcttaag tagctgaagc tccggttttg
     4021 aactatgcgc tcggggttgg cgagtgtgtt ttgtgaagtt ttttaggcac cttttgaaat
     4081 gtaatcattt gggtcaatat gtaattttca gtgttagact tgtaaattgt ccgctaaatt
     4141 ctggccgttt ttggcttttt tgttagacaa catgggtaaa aagcctgaac tcaccgcgac
     4201 gtctgtcgag aagtttctga tcgaaaagtt cgacagcgtc tccgacctga tgcagctctc
     4261 ggagggcgaa gaatctcgtg ctttcagctt cgatgtagga gggcgtggat atgtcctgcg
     4321 ggtaaatagc tgcgccgatg gtttctacaa agatcgttat gtttatcggc actttgcatc
     4381 cgccgcgctc ccgattccgg aagtgcttga cattggggag ttcagcgaga gcctgaccta
     4441 ttgcatctcc cgccgtgcac agggtgtcac gttgcaagac ctgcctgaaa ccgaactgcc
     4501 cgctgttctg cagccggtcg cggaggcaat ggatgccatc gctgccgccg atcttagcca
     4561 gacgagcggg ttcggcccat tcggaccgca aggaatcggt caatacacta catggcgtga
     4621 tttcatatgc gcgattgctg atccccatgt gtatcactgg caaactgtga tggacgacac
     4681 cgtcagtgcg tccgtcgcgc aggctctcga tgagctgatg ctttgggccg aggactgccc
     4741 cgaagtccgg cacctcgtgc acgcggattt cggctccaac aatgtcctga cggacaatgg
     4801 ccgcataaca gcggtcattg actggagcga ggcgatgttc ggggattccc aatacgaggt
     4861 cgccaacatc ttcttctgga ggccgtggtt ggcttgtatg gagcagcaga cgcgctactt
     4921 cgagcggagg catccggagc ttgcaggatc gccccggctc cgggcgtata tgctccgcat
     4981 tggtcttgac caactctatc agagcttggt tgacggcaat ttcgatgatg cagcttgggc
     5041 gcagggtcga tgcgacgcaa tcgtccgatc cggagccggg actgtcgggc gtacacaaat
     5101 cgcccgcaga agcgccgccg tctggaccga tggctgtgta gaagtactcg ccgatagtgg
     5161 aaaccgacgc cccagcactc gtccgagggc aaaggaataa gctagtatgt aagcctagtc
     5221 ttagataata aaatcgctat ccatcgaaga tggatgtgtg ttggtttttt gtgtgtgtaa
     5281 cgctaggcgc gcctggtgta ccgagaacga tcctctcagt gcgagtctcg acgatccata
     5341 tcgttgcttg gcagtcagcc agtcggaatc cagcttggga cccaggaagt ccaatcgtca
     5401 gatattgtac tcaagcctgg tcacggcagc gtaccgatct gtttaaacct agatattgat
     5461 agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc aaacatctat caagagacag
     5521 gatcagcagg aggctttcgc atgagtattc aacatttccg tgtcgccctt attccctttt
     5581 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     5641 ctgaagatca gttgggtgcg cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     5701 tccttgagag ttttcgcccc gaagaacgct ttccaatgat gagcactttt aaagttctgc
     5761 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     5821 actattctca gaatgacttg gttgagtatt caccagtcac agaaaagcat cttacggatg
     5881 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     5941 acttacttct gacaacgatt ggaggaccga aggagctaac cgcttttttg cacaacatgg
     6001 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     6061 acgagcgtga caccacgatg cctgtagcaa tggcaacaac cttgcgtaaa ctattaactg
     6121 gcgaactact tactctagct tcccggcaac agttgataga ctggatggag gcggataaag
     6181 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     6241 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     6301 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     6361 agatcgctga gataggtgcc tcactgatta agcattggta accgattcta ggtgcattgg
     6421 cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata ctcccacata tgccagattc
     6481 agcaacggat acggcttccc caacttgccc acttccatac gtgtcctcct taccagaaat
     6541 ttatccttaa gatcgtttaa actcgactct ggctctatcg aatctccgtc gtttcgagct
     6601 tacgcgaaca gccgtggcgc tcatttgctc gtcgggcatc gaatctcgtc agctatcgtc
     6661 agcttacctt tttggcagcg atcgcggctc ccgacatctt ggaccattag ctccacaggt
     6721 atcttcttcc ctctagtggt cataacagca gcttcagcta cctctcaatt caaaaaaccc
     6781 ctcaagaccc gtttagaggc cccaaggggt tatgctatca atcgttgcgt tacacacaca
     6841 aaaaaccaac acacatccat cttcgatgga tagcgatttt attatctaac tgctgatcga
     6901 gtgtagccag atctagtaat caattacggg gtcattagtt catagcccat atatggagtt
     6961 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
     7021 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
     7081 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
     7141 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
     7201 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
     7261 taccatgctg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
     7321 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
     7381 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
     7441 tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga tcagatcttt
     7501 gtcgatccta ccatccactc gacacacccg ccagcggccg c
//
