LOCUS       sequence-323979-        8096 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8096)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 8096)
  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-m4133362
FEATURES             Location/Qualifiers
     source          1..8096
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1916..1936
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1940..2005
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    2058..2179
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2069..2088)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2123..2142
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      2294..2340
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      2370..2506
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    2551..2622
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(2848..3436)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2928..2947)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        3936..5145
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     4982..5001
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             5149..5748
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(5149..5168)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5645..5665
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    5775..5823
                     /note="synthetic polyadenylation signal"
     CDS             join(6089..6157,6158..6949)
                     /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(6307..6326)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      6962..7098
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    7250..7321
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(7326..7372)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    7390..7438
                     /note="synthetic polyadenylation signal"
     enhancer        7519..7822
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        7824..8026
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     7977..7997
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgtac actcgctatg tcgataacaa cttctgtggc cctgatggct accctcttga
       61 gtgcattaaa gaccttctag cacgtgctgg taaagcttca tgcactttgt ccgaacaact
      121 ggactttatt gacactaaga ggggtgtata ctgctgccgt gaacatgagc atgaaattgc
      181 ttggtacacg gaacgttctg aaaagagcta tgaattgcag acaccttttg aaattaaatt
      241 ggcaaagaaa tttgacacct tcaatgggga atgtccaaat tttgtatttc ccttaaattc
      301 cataatcaag actattcaac caagggttga aaagaaaaag cttgatggct ttatgggtag
      361 aattcgatct gtctatccag ttgcgtcacc aaatgaatgc aaccaaatgt gcctttcaac
      421 tctcatgaag tgtgatcatt gtggtgaaac ttcatggcag acgggcgatt ttgttaaagc
      481 cacttgcgaa ttttgtggca ctgagaattt gactaaagaa ggtgccacta cttgtggtta
      541 cttaccccaa aatgctgttg ttaaaattta ttgtccagca tgtcacaatt cagaagtagg
      601 acctgagcat agtcttgccg aataccataa tgaatctggc ttgaaaacca ttcttcgtaa
      661 gggtggtcgc actattgcct ttggaggctg tgtgttctct tatgttggtt gccataacaa
      721 gtgtgcctat tgggttccac gtgctagcgc taacataggt tgtaaccata caggtgttgt
      781 tggagaaggt tccgaaggtc ttaatgacaa ccttcttgaa atactccaaa aagagaaagt
      841 caacatcaat attgttggtg actttaaact taatgaagag atcgccatta ttttggcatc
      901 tttttctgct tccacaagtg cttttgtgga aactgtgaaa ggtttggatt ataaagcatt
      961 caaacaaatt gttgaatcct gtggtaattt taaagttaca aaaggaaaag ctaaaaaagg
     1021 tgcctggaat attggtgaac agaaatcaat actgagtcct ctttatgcat ttgcatcaga
     1081 ggctgctcgt gttgtacgat caattttctc ccgcactctt gaaactgctc aaaattctgt
     1141 gcgtgtttta cagaaggccg ctataacaat actagatgga atttcacagt attcactgag
     1201 actcattgat gctatgatgt tcacatctga tttggctact aacaatctag ttgtaatggc
     1261 ctacattaca ggtggtgttg ttcagttgac ttcgcagtgg ctaactaaca tctttggcac
     1321 tgtttatgaa aaactcaaac ccgtccttga ttggcttgaa gagaagttta aggaaggtgt
     1381 agagtttctt agagacggtt gggaaattgt taaatttatc tcaacctgtg cttgtgaaat
     1441 tgtcggtgga caaattgtca cctgtgcaaa ggaaattaag gagagtgttc agacattctt
     1501 taagcttgta aataaatttt tggctttgtg tgctgactct atcattattg gtggagctaa
     1561 acttaaagcc ttgaatttag gtgaaacatt tgtcacgcac tcaaagggat tgtacagaaa
     1621 gtgtgttaaa tccagagaag aaactggcct actcatgcct ctaaaagccc caaaagaaat
     1681 tatcttctta gagggagaaa cacttcccac agaagtgtta acagaggaag ttgtcttgaa
     1741 aactggtgat ttacaaccat tagaacaacc tactagtgaa gctgttgaag ctccattggt
     1801 tggtacacca gtttgtatta acgggcttat gttgctcgaa atcaaagaca cagaaaagta
     1861 ctgtgccctt gcacctaata tgatggtaac aaacaatacc ttcacactca aaggcgaaaa
     1921 cctttacttt cagtcgatgg attataagga ccacgatgga gactacaaag atcatgacat
     1981 tgattacaag gacgacgatg acaaataagc tagcttgact gactgagata cagcgtacct
     2041 tcagctcaca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     2101 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     2161 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     2221 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta ttggcccatc
     2281 tctatcggta tcgtagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     2341 tgcccctcgg gccggattgc tatctaccgg cattggcgca gaaaaaaatg cctgatgcga
     2401 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac
     2461 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaaggtc gtcagctatc
     2521 ctgcaggcga tctctcgatt tcgatcaaga cattccttta atggtctttt ctggacacca
     2581 ctaggggtca gaagtagttc atcaaacttt cttccctccc taatctcatt ggttaccttg
     2641 ggctatcgaa acttaattaa ccagtcaagt cagctacttg gcgagatcga cttgtctggg
     2701 tttcgactac gctcagaatt gcgtcagtca agttcgatct ggtccttgct attgcacccg
     2761 ttctccgatt acgagtttca tttaaatcat gtgagcaaaa ggccagcaaa aggccaggaa
     2821 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     2881 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     2941 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     3001 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     3061 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     3121 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     3181 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     3241 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     3301 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3361 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     3421 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     3481 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     3541 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     3601 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     3661 atccatagtt gcatttaaat ttccgaactc tccaaggccc tcgtcggaaa atcttcaaac
     3721 ctttcgtccg atccatcttg caggctacct ctcgaacgaa ctatcgcaag tctcttggcc
     3781 ggccttgcgc cttggctatt gcttggcagc gcctatcgcc aggtattact ccaatcccga
     3841 atatccgaga tcgggatcac ccgagagaag ttcaacctac atcctcaatc ccgatctatc
     3901 cgagatccga ggaatatcga aatcggggcg cgcctggcct ccgcgccggg ttttggcgcc
     3961 tcccgcgggc gcccccctcg tcacggcgag cgctgccacg tcagacgaag ggcgcaggag
     4021 cgtcctgatc cttccgcccg gacgctcagg acagcggccc gctgctcata agactcggcc
     4081 ttagaacccc agtatcagca gaaggacatt ttaggacggg acttgggtga ctctagggca
     4141 ctggttttct ttccagagag cggaacaggc gaggaaaagt agtcccttct cggcgattct
     4201 gcggagggat ctccgtgggg cggtgaacgc cgatgattat ataaggacgc gccgggtgtg
     4261 gcacagctag ttccgtcgca gccgggattt gggtcgcggt tcttgtttgt ggatcgctgt
     4321 gatcgtcact tggtgagtag cgggctgctg ggctggccgg ggctttcgtg gccgccgggc
     4381 cgctcggtgg gacggaagcg tgtggagaga ccgccaaggg ctgtagtctg ggtccgcgag
     4441 caaggttgcc ctgaactggg ggttgggggg agcgcagcaa aatggcggct gttcccgagt
     4501 cttgaatgga agacgcttgt gaggcgggct gtgaggtcgt tgaaacaagg tggggggcat
     4561 ggtgggcggc aagaacccaa ggtcttgagc ccttcgctaa tgcgggaaag ctcttattcg
     4621 ggtgagatgg gctgggcacc atctggggac cctgacgtga agtttgtcac tgactggaga
     4681 actcggtttg tcgtctgttg cgggggcggc agttatggcg gtgccgttgg gcagtgcacc
     4741 cgtacctttg ggagcgcgcg ccctcgtcgt gtcgtgacgt cacccgttct gttggcttat
     4801 aatgcagggt ggggccacct gccggtaggt gtgcggtagg cttttctccg tcgcaggacg
     4861 cagggttcgg gcctagggta ggctctcctg aatcgacagg cgccggacct ctggtgaggg
     4921 gagggataag tgaggcgtca gtttctttgg tcggttttat gtacctatct tcttaagtag
     4981 ctgaagctcc ggttttgaac tatgcgctcg gggttggcga gtgtgttttg tgaagttttt
     5041 taggcacctt ttgaaatgta atcatttggg tcaatatgta attttcagtg ttagacttgt
     5101 aaattgtccg ctaaattctg gccgtttttg gcttttttgt tagacaacat gaccgagtac
     5161 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     5221 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     5281 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     5341 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     5401 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     5461 gcgcagcaac agatggaggg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     5521 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     5581 ctccccggag tggaggccgc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     5641 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     5701 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctaggc tagtatgtaa
     5761 gcctagtctt agataataaa atcgctatcc atcgaagatg gatgtgtgtt ggttttttgt
     5821 gtgtgtaacg ctaggcgcgc ctggtgtacc gagaacgatc ctctcagtgc gagtctcgac
     5881 gatccatatc gttgcttggc agtcagccag tcggaatcca gcttgggacc caggaagtcc
     5941 aatcgtcaga tattgtactc aagcctggtc acggcagcgt accgatctgt ttaaacctag
     6001 atattgatag tctgatcggt caacgtataa tcgagtccta gcttttgcaa acatctatca
     6061 agagacagga tcagcaggag gctttcgcat gagtattcaa catttccgtg tcgcccttat
     6121 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     6181 aaaagatgct gaagatcagt tgggtgcgcg agtgggttac atcgaactgg atctcaacag
     6241 cggtaagatc cttgagagtt ttcgccccga agaacgcttt ccaatgatga gcacttttaa
     6301 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     6361 ccgcatacac tattctcaga atgacttggt tgagtattca ccagtcacag aaaagcatct
     6421 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     6481 tgcggccaac ttacttctga caacgattgg aggaccgaag gagctaaccg cttttttgca
     6541 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     6601 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacct tgcgtaaact
     6661 attaactggc gaactactta ctctagcttc ccggcaacag ttgatagact ggatggaggc
     6721 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6781 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6841 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6901 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac cgattctagg
     6961 tgcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg
     7021 ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta
     7081 ccagaaattt atccttaaga tcccgaatcg tttaaactcg actctggctc tatcgaatct
     7141 ccgtcgtttc gagcttacgc gaacagccgt ggcgctcatt tgctcgtcgg gcatcgaatc
     7201 tcgtcagcta tcgtcagctt acctttttgg cagcgatcgc ggctcccgac atcttggacc
     7261 attagctcca caggtatctt cttccctcta gtggtcataa cagcagcttc agctacctct
     7321 caattcaaaa aacccctcaa gacccgttta gaggccccaa ggggttatgc tatcaatcgt
     7381 tgcgttacac acacaaaaaa ccaacacaca tccatcttcg atggatagcg attttattat
     7441 ctaactgctg atcgagtgta gccagatcta gtaatcaatt acggggtcat tagttcatag
     7501 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
     7561 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
     7621 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
     7681 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
     7741 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
     7801 attagtcatc gctattacca tgctgatgcg gttttggcag tacatcaatg ggcgtggata
     7861 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
     7921 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
     7981 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctggtt tagtgaaccg
     8041 tcagatcaga tctttgtcga tcctaccatc cactcgacac acccgccagc ggccgc
//
