LOCUS       sequence-295414-        7000 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 7000)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 7000)
  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-m4133205
FEATURES             Location/Qualifiers
     source          1..7000
                     /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             395..472
                     /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"
     CDS             500..520
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     polyA_signal    969..1090
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(980..999)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1034..1053
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      1205..1251
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      1281..1417
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    1462..1533
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1759..2347)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1839..1858)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2847..4056
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3893..3912
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             4060..4659
                     /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(4060..4079)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4556..4576
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4686..4734
                     /note="synthetic polyadenylation signal"
     CDS             join(5000..5068,5069..5860)
                     /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(5218..5237)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5873..6009
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    6154..6225
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(6230..6276)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    6294..6342
                     /note="synthetic polyadenylation signal"
     enhancer        6423..6726
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6728..6930
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6881..6901
                     /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 gggacggcga atcccggggc tgataaatcc ctggaaaagg agatggaaga agaactttat
      421 tgcagtgtct gcggctaacc ggttcaaaaa gatcagttca agcggcgccc tagattacga
      481 cattcctacg accgcctccg agaatctcta tttccagggt gaatttggct tggcacaaca
      541 tgacgaggcc gtcgataaca agtttaataa agaacagcag aacgctttct atgagatcct
      601 tcacttacca aatctgaacg aagagcaacg caacgctttc attcagagcc tgaaagacga
      661 tccctcgcag tctgcaaatc tcctggccga agccaagaag ctgaatgacg cacaggctcc
      721 aaaagttgat aacaagttta acaaggagca gcagaatgct ttctacgaga tattgcacct
      781 tcctaatctg aatgaggaac aaagaaatgc atttatccag tccctaaaag acgatccatc
      841 acagagtgcc aacctgcttg cggaagccaa aaagctcaat gatgcccagg ccccgaaagt
      901 agacgcgaac catcaataag ctagcttgac tgactgagat acagcgtacc ttcagctcac
      961 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     1021 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     1081 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg
     1141 ggaggttttt taaagcaagt aaaacctcta caaatgtggt attggcccat ctctatcggt
     1201 atcgtagcat aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gtgcccctcg
     1261 ggccggattg ctatctaccg gcattggcgc agaaaaaaat gcctgatgcg acgctgcgcg
     1321 tcttatactc ccacatatgc cagattcagc aacggatacg gcttccccaa cttgcccact
     1381 tccatacgtg tcctccttac cagaaattta tccttaaggt cgtcagctat cctgcaggcg
     1441 atctctcgat ttcgatcaag acattccttt aatggtcttt tctggacacc actaggggtc
     1501 agaagtagtt catcaaactt tcttccctcc ctaatctcat tggttacctt gggctatcga
     1561 aacttaatta accagtcaag tcagctactt ggcgagatcg acttgtctgg gtttcgacta
     1621 cgctcagaat tgcgtcagtc aagttcgatc tggtccttgc tattgcaccc gttctccgat
     1681 tacgagtttc atttaaatca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     1741 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     1801 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     1861 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     1921 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     1981 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     2041 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     2101 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     2161 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     2221 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     2281 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     2341 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     2401 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     2461 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     2521 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     2581 tgcatttaaa tttccgaact ctccaaggcc ctcgtcggaa aatcttcaaa cctttcgtcc
     2641 gatccatctt gcaggctacc tctcgaacga actatcgcaa gtctcttggc cggccttgcg
     2701 ccttggctat tgcttggcag cgcctatcgc caggtattac tccaatcccg aatatccgag
     2761 atcgggatca cccgagagaa gttcaaccta catcctcaat cccgatctat ccgagatccg
     2821 aggaatatcg aaatcggggc gcgcctggcc tccgcgccgg gttttggcgc ctcccgcggg
     2881 cgcccccctc gtcacggcga gcgctgccac gtcagacgaa gggcgcagga gcgtcctgat
     2941 ccttccgccc ggacgctcag gacagcggcc cgctgctcat aagactcggc cttagaaccc
     3001 cagtatcagc agaaggacat tttaggacgg gacttgggtg actctagggc actggttttc
     3061 tttccagaga gcggaacagg cgaggaaaag tagtcccttc tcggcgattc tgcggaggga
     3121 tctccgtggg gcggtgaacg ccgatgatta tataaggacg cgccgggtgt ggcacagcta
     3181 gttccgtcgc agccgggatt tgggtcgcgg ttcttgtttg tggatcgctg tgatcgtcac
     3241 ttggtgagta gcgggctgct gggctggccg gggctttcgt ggccgccggg ccgctcggtg
     3301 ggacggaagc gtgtggagag accgccaagg gctgtagtct gggtccgcga gcaaggttgc
     3361 cctgaactgg gggttggggg gagcgcagca aaatggcggc tgttcccgag tcttgaatgg
     3421 aagacgcttg tgaggcgggc tgtgaggtcg ttgaaacaag gtggggggca tggtgggcgg
     3481 caagaaccca aggtcttgag cccttcgcta atgcgggaaa gctcttattc gggtgagatg
     3541 ggctgggcac catctgggga ccctgacgtg aagtttgtca ctgactggag aactcggttt
     3601 gtcgtctgtt gcgggggcgg cagttatggc ggtgccgttg ggcagtgcac ccgtaccttt
     3661 gggagcgcgc gccctcgtcg tgtcgtgacg tcacccgttc tgttggctta taatgcaggg
     3721 tggggccacc tgccggtagg tgtgcggtag gcttttctcc gtcgcaggac gcagggttcg
     3781 ggcctagggt aggctctcct gaatcgacag gcgccggacc tctggtgagg ggagggataa
     3841 gtgaggcgtc agtttctttg gtcggtttta tgtacctatc ttcttaagta gctgaagctc
     3901 cggttttgaa ctatgcgctc ggggttggcg agtgtgtttt gtgaagtttt ttaggcacct
     3961 tttgaaatgt aatcatttgg gtcaatatgt aattttcagt gttagacttg taaattgtcc
     4021 gctaaattct ggccgttttt ggcttttttg ttagacaaca tgaccgagta caagcccacg
     4081 gtgcgcctcg ccacccgcga cgacgtcccc agggccgtac gcaccctcgc cgccgcgttc
     4141 gccgactacc ccgccacgcg ccacaccgtc gatccggacc gccacatcga gcgggtcacc
     4201 gagctgcaag aactcttcct cacgcgcgtc gggctcgaca tcggcaaggt gtgggtcgcg
     4261 gacgacggcg ccgcggtggc ggtctggacc acgccggaga gcgtcgaagc gggggcggtg
     4321 ttcgccgaga tcggcccgcg catggccgag ttgagcggtt cccggctggc cgcgcagcaa
     4381 cagatggagg gcctcctggc gccgcaccgg cccaaggagc ccgcgtggtt cctggccacc
     4441 gtcggcgtct cgcccgacca ccagggcaag ggtctgggca gcgccgtcgt gctccccgga
     4501 gtggaggccg ccgagcgcgc cggggtgccc gccttcctgg agacctccgc gccccgcaac
     4561 ctccccttct acgagcggct cggcttcacc gtcaccgccg acgtcgaggt gcccgaagga
     4621 ccgcgcacct ggtgcatgac ccgcaagccc ggtgcctagg ctagtatgta agcctagtct
     4681 tagataataa aatcgctatc catcgaagat ggatgtgtgt tggttttttg tgtgtgtaac
     4741 gctaggcgcg cctggtgtac cgagaacgat cctctcagtg cgagtctcga cgatccatat
     4801 cgttgcttgg cagtcagcca gtcggaatcc agcttgggac ccaggaagtc caatcgtcag
     4861 atattgtact caagcctggt cacggcagcg taccgatctg tttaaaccta gatattgata
     4921 gtctgatcgg tcaacgtata atcgagtcct agcttttgca aacatctatc aagagacagg
     4981 atcagcagga ggctttcgca tgagtattca acatttccgt gtcgccctta ttcccttttt
     5041 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5101 tgaagatcag ttgggtgcgc gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5161 ccttgagagt tttcgccccg aagaacgctt tccaatgatg agcactttta aagttctgct
     5221 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5281 ctattctcag aatgacttgg ttgagtattc accagtcaca gaaaagcatc ttacggatgg
     5341 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5401 cttacttctg acaacgattg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5461 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5521 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacc ttgcgtaaac tattaactgg
     5581 cgaactactt actctagctt cccggcaaca gttgatagac tggatggagg cggataaagt
     5641 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     5701 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     5761 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     5821 gatcgctgag ataggtgcct cactgattaa gcattggtaa ccgattctag gtgcattggc
     5881 gcagaaaaaa atgcctgatg cgacgctgcg cgtcttatac tcccacatat gccagattca
     5941 gcaacggata cggcttcccc aacttgccca cttccatacg tgtcctcctt accagaaatt
     6001 tatccttaag atcgtttaaa ctcgactctg gctctatcga atctccgtcg tttcgagctt
     6061 acgcgaacag ccgtggcgct catttgctcg tcgggcatcg aatctcgtca gctatcgtca
     6121 gcttaccttt ttggcagcga tcgcggctcc cgacatcttg gaccattagc tccacaggta
     6181 tcttcttccc tctagtggtc ataacagcag cttcagctac ctctcaattc aaaaaacccc
     6241 tcaagacccg tttagaggcc ccaaggggtt atgctatcaa tcgttgcgtt acacacacaa
     6301 aaaaccaaca cacatccatc ttcgatggat agcgatttta ttatctaact gctgatcgag
     6361 tgtagccaga tctagtaatc aattacgggg tcattagttc atagcccata tatggagttc
     6421 cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga cccccgccca
     6481 ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt ccattgacgt
     6541 caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt gtatcatatg
     6601 ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag
     6661 tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt catcgctatt
     6721 accatgctga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt tgactcacgg
     6781 ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca ccaaaatcaa
     6841 cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg cggtaggcgt
     6901 gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat cagatctttg
     6961 tcgatcctac catccactcg acacacccgc cagcggccgc
//
