LOCUS       sequence-295027-        8093 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 8093)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8093)
  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-m4133165
FEATURES             Location/Qualifiers
     source          1..8093
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1502..1522
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1523..1564
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     polyA_signal    1633..1754
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1644..1663)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1698..1717
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1869..1915
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1945..2081
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2126..2197
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2423..3011)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2503..2522)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3511..4720
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4557..4576
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4724..5752
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     polyA_signal    5779..5827
                     /note="synthetic polyadenylation signal"
     CDS             join(6093..6161,6162..6953)
                     /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(6311..6330)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6966..7102
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7247..7318
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(7323..7369)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7387..7435
                     /note="synthetic polyadenylation signal"
     enhancer        7516..7819
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7821..8023
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7974..7994
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgatt gttaataatt ggttgaagca gttaattaaa gttacacttg tgttcctttt
       61 tgttgctgct attttctatt taataacacc tgttcatgtc atgtctaaac atactgactt
      121 ttcaagtgaa atcataggat acaaggctat tgatggtggt gtcactcgtg acatagcatc
      181 tacagatact tgttttgcta acaaacatgc tgattttgac acatggttta gccagcgtgg
      241 tggtagttat actaatgaca aagcttgccc attgattgct gcagtcataa caagagaagt
      301 gggttttgtc gtgcctggtt tgcctggcac gatattacgc acaactaatg gtgacttttt
      361 gcatttctta cctagagttt ttagtgcagt tggtaacatc tgttacacac catcaaaact
      421 tatagagtac actgactttg caacatcagc ttgtgttttg gctgctgaat gtacaatttt
      481 taaagatgct tctggtaagc cagtaccata ttgttatgat accaatgtac tagaaggttc
      541 tgttgcttat gaaagtttac gccctgacac acgttatgtg ctcatggatg gctctattat
      601 tcaatttcct aacacctacc ttgaaggttc tgttagagtg gtaacaactt ttgattctga
      661 gtactgtagg cacggcactt gtgaaagatc agaagctggt gtttgtgtat ctactagtgg
      721 tagatgggta cttaacaatg attattacag atctttacca ggagttttct gtggtgtaga
      781 tgctgtaaat ttacttacta atatgtttac accactaatt caacctattg gtgctttgga
      841 catatcagca tctatagtag ctggtggtat tgtagctatc gtagtaacat gccttgccta
      901 ctattttatg aggtttagaa gagcttttgg tgaatacagt catgtagttg cctttaatac
      961 tttactattc cttatgtcat tcactgtact ctgtttaaca ccagtttact cattcttacc
     1021 tggtgtttat tctgttattt acttgtactt gacattttat cttactaatg atgtttcttt
     1081 tttagcacat attcagtgga tggttatgtt cacaccttta gtacctttct ggataacaat
     1141 tgcttatatc atttgtattt ccacaaagca tttctattgg ttctttagta attacctaaa
     1201 gagacgtgta gtctttaatg gtgtttcctt tagtactttt gaagaagctg cgctgtgcac
     1261 ctttttgtta aataaagaaa tgtatctaaa gttgcgtagt gatgtgctat tacctcttac
     1321 gcaatataat agatacttag ctctttataa taagtacaag tattttagtg gagcaatgga
     1381 tacaactagc tacagagaag ctgcttgttg tcatctcgca aaggctctca atgacttcag
     1441 taactcaggt tctgatgttc tttaccaacc accacaaacc tctatcacct cagctgtttt
     1501 ggaaaacctt tactttcagt cggggaaacc tatacccaat ccacttctcg gcttagattc
     1561 tacctaagga tccgcaggcc tctgctagct tgactgactg agatacagcg taccttcagc
     1621 tcacagacat gataagatac attgatgagt ttggacaaac cacaactaga atgcagtgaa
     1681 aaaaatgctt tatttgtgaa atttgtgatg ctattgcttt atttgtaacc attataagct
     1741 gcaataaaca agttaacaac aacaattgca ttcattttat gtttcaggtt cagggggagg
     1801 tgtgggaggt tttttaaagc aagtaaaacc tctacaaatg tggtattggc ccatctctat
     1861 cggtatcgta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgtgccc
     1921 ctcgggccgg attgctatct accggcattg gcgcagaaaa aaatgcctga tgcgacgctg
     1981 cgcgtcttat actcccacat atgccagatt cagcaacgga tacggcttcc ccaacttgcc
     2041 cacttccata cgtgtcctcc ttaccagaaa tttatcctta aggtcgtcag ctatcctgca
     2101 ggcgatctct cgatttcgat caagacattc ctttaatggt cttttctgga caccactagg
     2161 ggtcagaagt agttcatcaa actttcttcc ctccctaatc tcattggtta ccttgggcta
     2221 tcgaaactta attaaccagt caagtcagct acttggcgag atcgacttgt ctgggtttcg
     2281 actacgctca gaattgcgtc agtcaagttc gatctggtcc ttgctattgc acccgttctc
     2341 cgattacgag tttcatttaa atcatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2401 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2461 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2521 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2581 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2641 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2701 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2761 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2821 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2881 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2941 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     3001 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     3061 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3121 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     3181 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3241 tagttgcatt taaatttccg aactctccaa ggccctcgtc ggaaaatctt caaacctttc
     3301 gtccgatcca tcttgcaggc tacctctcga acgaactatc gcaagtctct tggccggcct
     3361 tgcgccttgg ctattgcttg gcagcgccta tcgccaggta ttactccaat cccgaatatc
     3421 cgagatcggg atcacccgag agaagttcaa cctacatcct caatcccgat ctatccgaga
     3481 tccgaggaat atcgaaatcg gggcgcgcct ggcctccgcg ccgggttttg gcgcctcccg
     3541 cgggcgcccc cctcgtcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgtcc
     3601 tgatccttcc gcccggacgc tcaggacagc ggcccgctgc tcataagact cggccttaga
     3661 accccagtat cagcagaagg acattttagg acgggacttg ggtgactcta gggcactggt
     3721 tttctttcca gagagcggaa caggcgagga aaagtagtcc cttctcggcg attctgcgga
     3781 gggatctccg tggggcggtg aacgccgatg attatataag gacgcgccgg gtgtggcaca
     3841 gctagttccg tcgcagccgg gatttgggtc gcggttcttg tttgtggatc gctgtgatcg
     3901 tcacttggtg agtagcgggc tgctgggctg gccggggctt tcgtggccgc cgggccgctc
     3961 ggtgggacgg aagcgtgtgg agagaccgcc aagggctgta gtctgggtcc gcgagcaagg
     4021 ttgccctgaa ctgggggttg gggggagcgc agcaaaatgg cggctgttcc cgagtcttga
     4081 atggaagacg cttgtgaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg
     4141 gcggcaagaa cccaaggtct tgagcccttc gctaatgcgg gaaagctctt attcgggtga
     4201 gatgggctgg gcaccatctg gggaccctga cgtgaagttt gtcactgact ggagaactcg
     4261 gtttgtcgtc tgttgcgggg gcggcagtta tggcggtgcc gttgggcagt gcacccgtac
     4321 ctttgggagc gcgcgccctc gtcgtgtcgt gacgtcaccc gttctgttgg cttataatgc
     4381 agggtggggc cacctgccgg taggtgtgcg gtaggctttt ctccgtcgca ggacgcaggg
     4441 ttcgggccta gggtaggctc tcctgaatcg acaggcgccg gacctctggt gaggggaggg
     4501 ataagtgagg cgtcagtttc tttggtcggt tttatgtacc tatcttctta agtagctgaa
     4561 gctccggttt tgaactatgc gctcggggtt ggcgagtgtg ttttgtgaag ttttttaggc
     4621 accttttgaa atgtaatcat ttgggtcaat atgtaatttt cagtgttaga cttgtaaatt
     4681 gtccgctaaa ttctggccgt ttttggcttt tttgttagac aacatgggta aaaagcctga
     4741 actcaccgcg acgtctgtcg agaagtttct gatcgaaaag ttcgacagcg tctccgacct
     4801 gatgcagctc tcggagggcg aagaatctcg tgctttcagc ttcgatgtag gagggcgtgg
     4861 atatgtcctg cgggtaaata gctgcgccga tggtttctac aaagatcgtt atgtttatcg
     4921 gcactttgca tccgccgcgc tcccgattcc ggaagtgctt gacattgggg agttcagcga
     4981 gagcctgacc tattgcatct cccgccgtgc acagggtgtc acgttgcaag acctgcctga
     5041 aaccgaactg cccgctgttc tgcagccggt cgcggaggca atggatgcca tcgctgccgc
     5101 cgatcttagc cagacgagcg ggttcggccc attcggaccg caaggaatcg gtcaatacac
     5161 tacatggcgt gatttcatat gcgcgattgc tgatccccat gtgtatcact ggcaaactgt
     5221 gatggacgac accgtcagtg cgtccgtcgc gcaggctctc gatgagctga tgctttgggc
     5281 cgaggactgc cccgaagtcc ggcacctcgt gcacgcggat ttcggctcca acaatgtcct
     5341 gacggacaat ggccgcataa cagcggtcat tgactggagc gaggcgatgt tcggggattc
     5401 ccaatacgag gtcgccaaca tcttcttctg gaggccgtgg ttggcttgta tggagcagca
     5461 gacgcgctac ttcgagcgga ggcatccgga gcttgcagga tcgccccggc tccgggcgta
     5521 tatgctccgc attggtcttg accaactcta tcagagcttg gttgacggca atttcgatga
     5581 tgcagcttgg gcgcagggtc gatgcgacgc aatcgtccga tccggagccg ggactgtcgg
     5641 gcgtacacaa atcgcccgca gaagcgccgc cgtctggacc gatggctgtg tagaagtact
     5701 cgccgatagt ggaaaccgac gccccagcac tcgtccgagg gcaaaggaat aagctagtat
     5761 gtaagcctag tcttagataa taaaatcgct atccatcgaa gatggatgtg tgttggtttt
     5821 ttgtgtgtgt aacgctaggc gcgcctggtg taccgagaac gatcctctca gtgcgagtct
     5881 cgacgatcca tatcgttgct tggcagtcag ccagtcggaa tccagcttgg gacccaggaa
     5941 gtccaatcgt cagatattgt actcaagcct ggtcacggca gcgtaccgat ctgtttaaac
     6001 ctagatattg atagtctgat cggtcaacgt ataatcgagt cctagctttt gcaaacatct
     6061 atcaagagac aggatcagca ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc
     6121 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     6181 aagtaaaaga tgctgaagat cagttgggtg cgcgagtggg ttacatcgaa ctggatctca
     6241 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ctttccaatg atgagcactt
     6301 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     6361 gtcgccgcat acactattct cagaatgact tggttgagta ttcaccagtc acagaaaagc
     6421 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     6481 acactgcggc caacttactt ctgacaacga ttggaggacc gaaggagcta accgcttttt
     6541 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6601 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca accttgcgta
     6661 aactattaac tggcgaacta cttactctag cttcccggca acagttgata gactggatgg
     6721 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6781 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6841 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6901 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taaccgattc
     6961 taggtgcatt ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca
     7021 tatgccagat tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc
     7081 cttaccagaa atttatcctt aagatcgttt aaactcgact ctggctctat cgaatctccg
     7141 tcgtttcgag cttacgcgaa cagccgtggc gctcatttgc tcgtcgggca tcgaatctcg
     7201 tcagctatcg tcagcttacc tttttggcag cgatcgcggc tcccgacatc ttggaccatt
     7261 agctccacag gtatcttctt ccctctagtg gtcataacag cagcttcagc tacctctcaa
     7321 ttcaaaaaac ccctcaagac ccgtttagag gccccaaggg gttatgctat caatcgttgc
     7381 gttacacaca caaaaaacca acacacatcc atcttcgatg gatagcgatt ttattatcta
     7441 actgctgatc gagtgtagcc agatctagta atcaattacg gggtcattag ttcatagccc
     7501 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     7561 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     7621 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     7681 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     7741 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     7801 agtcatcgct attaccatgc tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
     7861 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     7921 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
     7981 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca
     8041 gatcagatct ttgtcgatcc taccatccac tcgacacacc cgccagcggc cgc
//
