LOCUS       sequence-295810-        8084 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp4.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8084)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8084)
  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-m4134398
FEATURES             Location/Qualifiers
     source          1..8084
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1508..1528
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1529..1555
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    1624..1745
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1635..1654)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1689..1708
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1860..1906
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1936..2072
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2117..2188
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2414..3002)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2494..2513)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3502..4711
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4548..4567
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4715..5743
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5770..5818
                     /note="synthetic polyadenylation signal"
     CDS             join(6084..6152,6153..6944)
                     /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(6302..6321)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6957..7093
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7238..7309
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(7314..7360)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7378..7426
                     /note="synthetic polyadenylation signal"
     enhancer        7507..7810
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7812..8014
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7965..7985
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgaag attgtaaata actggttgaa acaactaata aaggttactt tagtgttcct
       61 gtttgtcgcc gcaatcttct atcttatcac cccggtgcac gtgatgagta aacatacaga
      121 cttttcttca gaaattatcg ggtacaaggc tatcgatggc ggtgtaacgc gcgacattgc
      181 ctccacagat acctgcttcg cgaacaaaca cgctgacttt gatacttggt tcagccagcg
      241 aggaggctcg tacaccaatg acaaagcctg tccactgatc gcagccgtga taactagaga
      301 ggttggattc gtcgtccctg ggctccccgg gacaattctc cgtacaacca acggcgactt
      361 tctgcatttt ctgccccggg tgttctccgc agtcggtaat atctgttata ctccatctaa
      421 gctcattgag tacacggatt ttgctaccag cgcgtgcgtg ttggccgcag aatgcaccat
      481 atttaaggac gcttctggaa aacctgtgcc ctattgttac gataccaatg tgcttgaggg
      541 ctcagttgcc tatgaaagtc tgaggccaga cactagatac gttttaatgg atggaagcat
      601 catccagttc ccaaacacgt atctggaggg gtccgtgcgg gtcgtgacca cattcgattc
      661 agaatactgc cgccacggta catgcgagag gtctgaggct ggcgtatgtg tgtcaactag
      721 cggtcgatgg gttctaaata acgattatta tcgttctctg cccggcgttt tttgtggagt
      781 ggacgccgtt aaccttttga ccaacatgtt tacacctctc attcagccca ttggtgcact
      841 ggatatctcc gcgagtattg tggctggggg aatagtcgcc attgtcgtaa cctgcttggc
      901 atactatttc atgcgtttca ggcgagcttt tggcgaatac tcacatgtcg tggccttcaa
      961 tactcttctg tttctgatga gcttcacggt actctgtctg accccagtat acagtttcct
     1021 acctggggtg tattctgtga tctacttata cctcacattt tacctgacca atgacgtgtc
     1081 ctttctggcc cacatccaat ggatggttat gttcacgccg ctcgtgcctt tctggattac
     1141 aatcgcttat ataatctgta ttagcacgaa gcatttctat tggttctttt ccaactactt
     1201 aaagcggcgc gtcgtattta atggggtatc gtttagcact tttgaggaag ccgcgctctg
     1261 cactttcttg cttaataagg agatgtattt gaaattaaga tctgacgttc tgcttccttt
     1321 aacacagtac aacagatacc tggcgttgta taacaaatac aaatatttct cgggagccat
     1381 ggatacaact tcctatagag aggctgcttg ttgccacctt gcaaaggcac tcaacgactt
     1441 cagtaacagc ggcagtgatg tgctttatca gccccctcag acttcaataa catccgccgt
     1501 cctccaagaa aacctttact ttcagtcgta cccctatgac gtaccagatt atgcgtaagg
     1561 atccgcaggc ctctgctagc ttgactgact gagatacagc gtaccttcag ctcacagaca
     1621 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     1681 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     1741 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg
     1801 ttttttaaag caagtaaaac ctctacaaat gtggtattgg cccatctcta tcggtatcgt
     1861 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgtgcc cctcgggccg
     1921 gattgctatc taccggcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta
     1981 tactcccaca tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat
     2041 acgtgtcctc cttaccagaa atttatcctt aaggtcgtca gctatcctgc aggcgatctc
     2101 tcgatttcga tcaagacatt cctttaatgg tcttttctgg acaccactag gggtcagaag
     2161 tagttcatca aactttcttc cctccctaat ctcattggtt accttgggct atcgaaactt
     2221 aattaaccag tcaagtcagc tacttggcga gatcgacttg tctgggtttc gactacgctc
     2281 agaattgcgt cagtcaagtt cgatctggtc cttgctattg cacccgttct ccgattacga
     2341 gtttcattta aatcatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     2401 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     2461 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     2521 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     2581 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     2641 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     2701 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     2761 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     2821 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     2881 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     2941 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     3001 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     3061 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     3121 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     3181 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcat
     3241 ttaaatttcc gaactctcca aggccctcgt cggaaaatct tcaaaccttt cgtccgatcc
     3301 atcttgcagg ctacctctcg aacgaactat cgcaagtctc ttggccggcc ttgcgccttg
     3361 gctattgctt ggcagcgcct atcgccaggt attactccaa tcccgaatat ccgagatcgg
     3421 gatcacccga gagaagttca acctacatcc tcaatcccga tctatccgag atccgaggaa
     3481 tatcgaaatc ggggcgcgcc tggcctccgc gccgggtttt ggcgcctccc gcgggcgccc
     3541 ccctcgtcac ggcgagcgct gccacgtcag acgaagggcg caggagcgtc ctgatccttc
     3601 cgcccggacg ctcaggacag cggcccgctg ctcataagac tcggccttag aaccccagta
     3661 tcagcagaag gacattttag gacgggactt gggtgactct agggcactgg ttttctttcc
     3721 agagagcgga acaggcgagg aaaagtagtc ccttctcggc gattctgcgg agggatctcc
     3781 gtggggcggt gaacgccgat gattatataa ggacgcgccg ggtgtggcac agctagttcc
     3841 gtcgcagccg ggatttgggt cgcggttctt gtttgtggat cgctgtgatc gtcacttggt
     3901 gagtagcggg ctgctgggct ggccggggct ttcgtggccg ccgggccgct cggtgggacg
     3961 gaagcgtgtg gagagaccgc caagggctgt agtctgggtc cgcgagcaag gttgccctga
     4021 actgggggtt ggggggagcg cagcaaaatg gcggctgttc ccgagtcttg aatggaagac
     4081 gcttgtgagg cgggctgtga ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga
     4141 acccaaggtc ttgagccctt cgctaatgcg ggaaagctct tattcgggtg agatgggctg
     4201 ggcaccatct ggggaccctg acgtgaagtt tgtcactgac tggagaactc ggtttgtcgt
     4261 ctgttgcggg ggcggcagtt atggcggtgc cgttgggcag tgcacccgta cctttgggag
     4321 cgcgcgccct cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg cagggtgggg
     4381 ccacctgccg gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg gttcgggcct
     4441 agggtaggct ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg gataagtgag
     4501 gcgtcagttt ctttggtcgg ttttatgtac ctatcttctt aagtagctga agctccggtt
     4561 ttgaactatg cgctcggggt tggcgagtgt gttttgtgaa gttttttagg caccttttga
     4621 aatgtaatca tttgggtcaa tatgtaattt tcagtgttag acttgtaaat tgtccgctaa
     4681 attctggccg tttttggctt ttttgttaga caacatgggt aaaaagcctg aactcaccgc
     4741 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     4801 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     4861 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     4921 atccgccgcg ctcccgattc cggaagtgct tgacattggg gagttcagcg agagcctgac
     4981 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     5041 gcccgctgtt ctgcagccgg tcgcggaggc aatggatgcc atcgctgccg ccgatcttag
     5101 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     5161 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     5221 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     5281 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     5341 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     5401 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     5461 cttcgagcgg aggcatccgg agcttgcagg atcgccccgg ctccgggcgt atatgctccg
     5521 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     5581 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     5641 aatcgcccgc agaagcgccg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     5701 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa taagctagta tgtaagccta
     5761 gtcttagata ataaaatcgc tatccatcga agatggatgt gtgttggttt tttgtgtgtg
     5821 taacgctagg cgcgcctggt gtaccgagaa cgatcctctc agtgcgagtc tcgacgatcc
     5881 atatcgttgc ttggcagtca gccagtcgga atccagcttg ggacccagga agtccaatcg
     5941 tcagatattg tactcaagcc tggtcacggc agcgtaccga tctgtttaaa cctagatatt
     6001 gatagtctga tcggtcaacg tataatcgag tcctagcttt tgcaaacatc tatcaagaga
     6061 caggatcagc aggaggcttt cgcatgagta ttcaacattt ccgtgtcgcc cttattccct
     6121 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     6181 atgctgaaga tcagttgggt gcgcgagtgg gttacatcga actggatctc aacagcggta
     6241 agatccttga gagttttcgc cccgaagaac gctttccaat gatgagcact tttaaagttc
     6301 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     6361 tacactattc tcagaatgac ttggttgagt attcaccagt cacagaaaag catcttacgg
     6421 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     6481 ccaacttact tctgacaacg attggaggac cgaaggagct aaccgctttt ttgcacaaca
     6541 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     6601 acgacgagcg tgacaccacg atgcctgtag caatggcaac aaccttgcgt aaactattaa
     6661 ctggcgaact acttactcta gcttcccggc aacagttgat agactggatg gaggcggata
     6721 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     6781 ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     6841 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     6901 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaaccgatt ctaggtgcat
     6961 tggcgcagaa aaaaatgcct gatgcgacgc tgcgcgtctt atactcccac atatgccaga
     7021 ttcagcaacg gatacggctt ccccaacttg cccacttcca tacgtgtcct ccttaccaga
     7081 aatttatcct taagatcgtt taaactcgac tctggctcta tcgaatctcc gtcgtttcga
     7141 gcttacgcga acagccgtgg cgctcatttg ctcgtcgggc atcgaatctc gtcagctatc
     7201 gtcagcttac ctttttggca gcgatcgcgg ctcccgacat cttggaccat tagctccaca
     7261 ggtatcttct tccctctagt ggtcataaca gcagcttcag ctacctctca attcaaaaaa
     7321 cccctcaaga cccgtttaga ggccccaagg ggttatgcta tcaatcgttg cgttacacac
     7381 acaaaaaacc aacacacatc catcttcgat ggatagcgat tttattatct aactgctgat
     7441 cgagtgtagc cagatctagt aatcaattac ggggtcatta gttcatagcc catatatgga
     7501 gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg
     7561 cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg
     7621 acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca
     7681 tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc
     7741 ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc
     7801 tattaccatg ctgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
     7861 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
     7921 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
     7981 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatcagatc
     8041 tttgtcgatc ctaccatcca ctcgacacac ccgccagcgg ccgc
//
