LOCUS       sequence-295741-        6529 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for SARS-CoV-2 nsp9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6529)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6529)
  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-m4133912
FEATURES             Location/Qualifiers
     source          1..6529
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             347..367
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             368..445
                     /codon_start=1
                     /product="calmodulin-binding peptide"
                     /label=CBP
                     /note="derived from skeletal muscle myosin light chain 
                     kinase; binds calmodulin with nanomolar affinity in the 
                     presence of calcium"
                     /translation="KRRWKKNFIAVSAANRFKKISSSGAL"
     polyA_signal    498..619
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(509..528)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     563..582
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      734..780
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      810..946
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    991..1062
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1288..1876)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1368..1387)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2376..3585
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3422..3441
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3589..4188
                     /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(3589..3608)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4085..4105
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4215..4263
                     /note="synthetic polyadenylation signal"
     CDS             join(4529..4597,4598..5389)
                     /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(4747..4766)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5402..5538
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5683..5754
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5759..5805)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5823..5871
                     /note="synthetic polyadenylation signal"
     enhancer        5952..6255
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6257..6459
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6410..6430
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgaat aacgagttaa gcccagtagc gcttcgccag atgagttgcg ccgcaggtac
       61 aacccaaact gcttgtacgg atgacaacgc cctcgcttac tataatacca caaagggagg
      121 ccgattcgtt ctggccctgt tgtcggatct gcaagacttg aaatgggcac gttttcccaa
      181 gtccgacggg actggcacca tttacacaga actagagccg ccatgcaggt tcgtgactga
      241 tacccctaaa ggacccaagg tcaaatatct gtattttatc aaggggctta acaatctcaa
      301 tagaggcatg gtgttaggat ctctagccgc aacggtgcgg ctgcaagaaa acctttactt
      361 tcagtcgaag aggcgatgga aaaagaattt tatcgcggtg agtgcagcca acagattcaa
      421 aaagatatcc tcgtctggtg ccctctaagc tagcttgact gactgagata cagcgtacct
      481 tcagctcaca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
      541 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
      601 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
      661 ggaggtgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta ttggcccatc
      721 tctatcggta tcgtagcata accccttggg gcctctaaac gggtcttgag gggttttttg
      781 tgcccctcgg gccggattgc tatctaccgg cattggcgca gaaaaaaatg cctgatgcga
      841 cgctgcgcgt cttatactcc cacatatgcc agattcagca acggatacgg cttccccaac
      901 ttgcccactt ccatacgtgt cctccttacc agaaatttat ccttaaggtc gtcagctatc
      961 ctgcaggcga tctctcgatt tcgatcaaga cattccttta atggtctttt ctggacacca
     1021 ctaggggtca gaagtagttc atcaaacttt cttccctccc taatctcatt ggttaccttg
     1081 ggctatcgaa acttaattaa ccagtcaagt cagctacttg gcgagatcga cttgtctggg
     1141 tttcgactac gctcagaatt gcgtcagtca agttcgatct ggtccttgct attgcacccg
     1201 ttctccgatt acgagtttca tttaaatcat gtgagcaaaa ggccagcaaa aggccaggaa
     1261 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     1321 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     1381 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     1441 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     1501 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     1561 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     1621 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     1681 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     1741 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     1801 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     1861 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     1921 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     1981 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     2041 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     2101 atccatagtt gcatttaaat ttccgaactc tccaaggccc tcgtcggaaa atcttcaaac
     2161 ctttcgtccg atccatcttg caggctacct ctcgaacgaa ctatcgcaag tctcttggcc
     2221 ggccttgcgc cttggctatt gcttggcagc gcctatcgcc aggtattact ccaatcccga
     2281 atatccgaga tcgggatcac ccgagagaag ttcaacctac atcctcaatc ccgatctatc
     2341 cgagatccga ggaatatcga aatcggggcg cgcctggcct ccgcgccggg ttttggcgcc
     2401 tcccgcgggc gcccccctcg tcacggcgag cgctgccacg tcagacgaag ggcgcaggag
     2461 cgtcctgatc cttccgcccg gacgctcagg acagcggccc gctgctcata agactcggcc
     2521 ttagaacccc agtatcagca gaaggacatt ttaggacggg acttgggtga ctctagggca
     2581 ctggttttct ttccagagag cggaacaggc gaggaaaagt agtcccttct cggcgattct
     2641 gcggagggat ctccgtgggg cggtgaacgc cgatgattat ataaggacgc gccgggtgtg
     2701 gcacagctag ttccgtcgca gccgggattt gggtcgcggt tcttgtttgt ggatcgctgt
     2761 gatcgtcact tggtgagtag cgggctgctg ggctggccgg ggctttcgtg gccgccgggc
     2821 cgctcggtgg gacggaagcg tgtggagaga ccgccaaggg ctgtagtctg ggtccgcgag
     2881 caaggttgcc ctgaactggg ggttgggggg agcgcagcaa aatggcggct gttcccgagt
     2941 cttgaatgga agacgcttgt gaggcgggct gtgaggtcgt tgaaacaagg tggggggcat
     3001 ggtgggcggc aagaacccaa ggtcttgagc ccttcgctaa tgcgggaaag ctcttattcg
     3061 ggtgagatgg gctgggcacc atctggggac cctgacgtga agtttgtcac tgactggaga
     3121 actcggtttg tcgtctgttg cgggggcggc agttatggcg gtgccgttgg gcagtgcacc
     3181 cgtacctttg ggagcgcgcg ccctcgtcgt gtcgtgacgt cacccgttct gttggcttat
     3241 aatgcagggt ggggccacct gccggtaggt gtgcggtagg cttttctccg tcgcaggacg
     3301 cagggttcgg gcctagggta ggctctcctg aatcgacagg cgccggacct ctggtgaggg
     3361 gagggataag tgaggcgtca gtttctttgg tcggttttat gtacctatct tcttaagtag
     3421 ctgaagctcc ggttttgaac tatgcgctcg gggttggcga gtgtgttttg tgaagttttt
     3481 taggcacctt ttgaaatgta atcatttggg tcaatatgta attttcagtg ttagacttgt
     3541 aaattgtccg ctaaattctg gccgtttttg gcttttttgt tagacaacat gaccgagtac
     3601 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc
     3661 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag
     3721 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg
     3781 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg
     3841 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc
     3901 gcgcagcaac agatggaggg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc
     3961 ctggccaccg tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg
     4021 ctccccggag tggaggccgc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg
     4081 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg
     4141 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctaggc tagtatgtaa
     4201 gcctagtctt agataataaa atcgctatcc atcgaagatg gatgtgtgtt ggttttttgt
     4261 gtgtgtaacg ctaggcgcgc ctggtgtacc gagaacgatc ctctcagtgc gagtctcgac
     4321 gatccatatc gttgcttggc agtcagccag tcggaatcca gcttgggacc caggaagtcc
     4381 aatcgtcaga tattgtactc aagcctggtc acggcagcgt accgatctgt ttaaacctag
     4441 atattgatag tctgatcggt caacgtataa tcgagtccta gcttttgcaa acatctatca
     4501 agagacagga tcagcaggag gctttcgcat gagtattcaa catttccgtg tcgcccttat
     4561 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4621 aaaagatgct gaagatcagt tgggtgcgcg agtgggttac atcgaactgg atctcaacag
     4681 cggtaagatc cttgagagtt ttcgccccga agaacgcttt ccaatgatga gcacttttaa
     4741 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     4801 ccgcatacac tattctcaga atgacttggt tgagtattca ccagtcacag aaaagcatct
     4861 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     4921 tgcggccaac ttacttctga caacgattgg aggaccgaag gagctaaccg cttttttgca
     4981 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5041 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacct tgcgtaaact
     5101 attaactggc gaactactta ctctagcttc ccggcaacag ttgatagact ggatggaggc
     5161 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     5221 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     5281 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5341 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac cgattctagg
     5401 tgcattggcg cagaaaaaaa tgcctgatgc gacgctgcgc gtcttatact cccacatatg
     5461 ccagattcag caacggatac ggcttcccca acttgcccac ttccatacgt gtcctcctta
     5521 ccagaaattt atccttaaga tcgtttaaac tcgactctgg ctctatcgaa tctccgtcgt
     5581 ttcgagctta cgcgaacagc cgtggcgctc atttgctcgt cgggcatcga atctcgtcag
     5641 ctatcgtcag cttacctttt tggcagcgat cgcggctccc gacatcttgg accattagct
     5701 ccacaggtat cttcttccct ctagtggtca taacagcagc ttcagctacc tctcaattca
     5761 aaaaacccct caagacccgt ttagaggccc caaggggtta tgctatcaat cgttgcgtta
     5821 cacacacaaa aaaccaacac acatccatct tcgatggata gcgattttat tatctaactg
     5881 ctgatcgagt gtagccagat ctagtaatca attacggggt cattagttca tagcccatat
     5941 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     6001 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     6061 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     6121 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     6181 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     6241 atcgctatta ccatgctgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     6301 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     6361 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     6421 ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa ccgtcagatc
     6481 agatctttgt cgatcctacc atccactcga cacacccgcc agcggccgc
//
