LOCUS       sequence-295446-        6463 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 6463)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6463)
  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-m4133986
FEATURES             Location/Qualifiers
     source          1..6463
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             341..361
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             362..379
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    432..553
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(443..462)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     497..516
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      668..714
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      744..880
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    925..996
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1222..1810)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1302..1321)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2310..3519
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3356..3375
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3523..4122
                     /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(3523..3542)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4019..4039
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4149..4197
                     /note="synthetic polyadenylation signal"
     CDS             join(4463..4531,4532..5323)
                     /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(4681..4700)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5336..5472
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5617..5688
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5693..5739)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5757..5805
                     /note="synthetic polyadenylation signal"
     enhancer        5886..6189
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6191..6393
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6344..6364
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 caccatgaat gagcttagtc ctgttgcact acgacagatg tcttgtgctg ccggtactac
       61 acaaactgct tgcactgatg acaatgcgtt agcttactac aacacaacaa agggaggtag
      121 gtttgtactt gcactgttat ccgatttaca ggatttgaaa tgggctagat tccctaagag
      181 tgatggaact ggtactatct atacagaact ggaaccacct tgtaggtttg ttacagacac
      241 acctaaaggt cctaaagtga agtatttata ctttattaaa ggattaaaca acctaaatag
      301 aggtatggta cttggtagtt tagctgccac agtacgtcta gaaaaccttt actttcagtc
      361 gcaccatcac catcaccatt aagctagctt gactgactga gatacagcgt accttcagct
      421 cacagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa
      481 aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg
      541 caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc agggggaggt
      601 gtgggaggtt ttttaaagca agtaaaacct ctacaaatgt ggtattggcc catctctatc
      661 ggtatcgtag cataacccct tggggcctct aaacgggtct tgaggggttt tttgtgcccc
      721 tcgggccgga ttgctatcta ccggcattgg cgcagaaaaa aatgcctgat gcgacgctgc
      781 gcgtcttata ctcccacata tgccagattc agcaacggat acggcttccc caacttgccc
      841 acttccatac gtgtcctcct taccagaaat ttatccttaa ggtcgtcagc tatcctgcag
      901 gcgatctctc gatttcgatc aagacattcc tttaatggtc ttttctggac accactaggg
      961 gtcagaagta gttcatcaaa ctttcttccc tccctaatct cattggttac cttgggctat
     1021 cgaaacttaa ttaaccagtc aagtcagcta cttggcgaga tcgacttgtc tgggtttcga
     1081 ctacgctcag aattgcgtca gtcaagttcg atctggtcct tgctattgca cccgttctcc
     1141 gattacgagt ttcatttaaa tcatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     1201 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     1261 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     1321 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     1381 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     1441 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     1501 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     1561 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     1621 agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg
     1681 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     1741 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     1801 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     1861 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     1921 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     1981 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     2041 agttgcattt aaatttccga actctccaag gccctcgtcg gaaaatcttc aaacctttcg
     2101 tccgatccat cttgcaggct acctctcgaa cgaactatcg caagtctctt ggccggcctt
     2161 gcgccttggc tattgcttgg cagcgcctat cgccaggtat tactccaatc ccgaatatcc
     2221 gagatcggga tcacccgaga gaagttcaac ctacatcctc aatcccgatc tatccgagat
     2281 ccgaggaata tcgaaatcgg ggcgcgcctg gcctccgcgc cgggttttgg cgcctcccgc
     2341 gggcgccccc ctcgtcacgg cgagcgctgc cacgtcagac gaagggcgca ggagcgtcct
     2401 gatccttccg cccggacgct caggacagcg gcccgctgct cataagactc ggccttagaa
     2461 ccccagtatc agcagaagga cattttagga cgggacttgg gtgactctag ggcactggtt
     2521 ttctttccag agagcggaac aggcgaggaa aagtagtccc ttctcggcga ttctgcggag
     2581 ggatctccgt ggggcggtga acgccgatga ttatataagg acgcgccggg tgtggcacag
     2641 ctagttccgt cgcagccggg atttgggtcg cggttcttgt ttgtggatcg ctgtgatcgt
     2701 cacttggtga gtagcgggct gctgggctgg ccggggcttt cgtggccgcc gggccgctcg
     2761 gtgggacgga agcgtgtgga gagaccgcca agggctgtag tctgggtccg cgagcaaggt
     2821 tgccctgaac tgggggttgg ggggagcgca gcaaaatggc ggctgttccc gagtcttgaa
     2881 tggaagacgc ttgtgaggcg ggctgtgagg tcgttgaaac aaggtggggg gcatggtggg
     2941 cggcaagaac ccaaggtctt gagcccttcg ctaatgcggg aaagctctta ttcgggtgag
     3001 atgggctggg caccatctgg ggaccctgac gtgaagtttg tcactgactg gagaactcgg
     3061 tttgtcgtct gttgcggggg cggcagttat ggcggtgccg ttgggcagtg cacccgtacc
     3121 tttgggagcg cgcgccctcg tcgtgtcgtg acgtcacccg ttctgttggc ttataatgca
     3181 gggtggggcc acctgccggt aggtgtgcgg taggcttttc tccgtcgcag gacgcagggt
     3241 tcgggcctag ggtaggctct cctgaatcga caggcgccgg acctctggtg aggggaggga
     3301 taagtgaggc gtcagtttct ttggtcggtt ttatgtacct atcttcttaa gtagctgaag
     3361 ctccggtttt gaactatgcg ctcggggttg gcgagtgtgt tttgtgaagt tttttaggca
     3421 ccttttgaaa tgtaatcatt tgggtcaata tgtaattttc agtgttagac ttgtaaattg
     3481 tccgctaaat tctggccgtt tttggctttt ttgttagaca acatgaccga gtacaagccc
     3541 acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct cgccgccgcg
     3601 ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat cgagcgggtc
     3661 accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc
     3721 gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga agcgggggcg
     3781 gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct ggccgcgcag
     3841 caacagatgg agggcctcct ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc
     3901 accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc
     3961 ggagtggagg ccgccgagcg cgccggggtg cccgccttcc tggagacctc cgcgccccgc
     4021 aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa
     4081 ggaccgcgca cctggtgcat gacccgcaag cccggtgcct aggctagtat gtaagcctag
     4141 tcttagataa taaaatcgct atccatcgaa gatggatgtg tgttggtttt ttgtgtgtgt
     4201 aacgctaggc gcgcctggtg taccgagaac gatcctctca gtgcgagtct cgacgatcca
     4261 tatcgttgct tggcagtcag ccagtcggaa tccagcttgg gacccaggaa gtccaatcgt
     4321 cagatattgt actcaagcct ggtcacggca gcgtaccgat ctgtttaaac ctagatattg
     4381 atagtctgat cggtcaacgt ataatcgagt cctagctttt gcaaacatct atcaagagac
     4441 aggatcagca ggaggctttc gcatgagtat tcaacatttc cgtgtcgccc ttattccctt
     4501 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     4561 tgctgaagat cagttgggtg cgcgagtggg ttacatcgaa ctggatctca acagcggtaa
     4621 gatccttgag agttttcgcc ccgaagaacg ctttccaatg atgagcactt ttaaagttct
     4681 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     4741 acactattct cagaatgact tggttgagta ttcaccagtc acagaaaagc atcttacgga
     4801 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     4861 caacttactt ctgacaacga ttggaggacc gaaggagcta accgcttttt tgcacaacat
     4921 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     4981 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca accttgcgta aactattaac
     5041 tggcgaacta cttactctag cttcccggca acagttgata gactggatgg aggcggataa
     5101 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     5161 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     5221 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     5281 acagatcgct gagataggtg cctcactgat taagcattgg taaccgattc taggtgcatt
     5341 ggcgcagaaa aaaatgcctg atgcgacgct gcgcgtctta tactcccaca tatgccagat
     5401 tcagcaacgg atacggcttc cccaacttgc ccacttccat acgtgtcctc cttaccagaa
     5461 atttatcctt aagatcgttt aaactcgact ctggctctat cgaatctccg tcgtttcgag
     5521 cttacgcgaa cagccgtggc gctcatttgc tcgtcgggca tcgaatctcg tcagctatcg
     5581 tcagcttacc tttttggcag cgatcgcggc tcccgacatc ttggaccatt agctccacag
     5641 gtatcttctt ccctctagtg gtcataacag cagcttcagc tacctctcaa ttcaaaaaac
     5701 ccctcaagac ccgtttagag gccccaaggg gttatgctat caatcgttgc gttacacaca
     5761 caaaaaacca acacacatcc atcttcgatg gatagcgatt ttattatcta actgctgatc
     5821 gagtgtagcc agatctagta atcaattacg gggtcattag ttcatagccc atatatggag
     5881 ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc
     5941 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
     6001 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
     6061 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
     6121 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
     6181 attaccatgc tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca
     6241 cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat
     6301 caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg
     6361 cgtgtacggt gggaggtcta tataagcaga gctggtttag tgaaccgtca gatcagatct
     6421 ttgtcgatcc taccatccac tcgacacacc cgccagcggc cgc
//
