LOCUS       sequence-295437-        6490 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 6490)
  TITLE     SARS-CoV-2 protein Mammalian expression constructs, tagged, for 
            protein expression and/or purification
REFERENCE   2  (bases 1 to 6490)
  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-m4133998
FEATURES             Location/Qualifiers
     source          1..6490
                     /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..406
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     polyA_signal    459..580
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(470..489)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     524..543
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     terminator      695..741
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     terminator      771..907
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    952..1023
                     /label=3' beta-globin insulator
                     /note="insulator downstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     rep_origin      complement(1249..1837)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1329..1348)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2337..3546
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3383..3402
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     CDS             3550..4149
                     /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(3550..3569)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     4046..4066
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     polyA_signal    4176..4224
                     /note="synthetic polyadenylation signal"
     CDS             join(4490..4558,4559..5350)
                     /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(4708..4727)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      5363..5499
                     /label=rrnG terminator
                     /note="transcription terminator from the E. coli ribosomal 
                     RNA rrnG operon (Albrechtsen et al., 1991)"
     misc_feature    5644..5715
                     /label=5' beta-globin insulator
                     /note="insulator upstream of the human beta-globin locus 
                     (Farrell et al., 2002)"
     terminator      complement(5720..5766)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     polyA_signal    5784..5832
                     /note="synthetic polyadenylation signal"
     enhancer        5913..6216
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6218..6420
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6371..6391
                     /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 gaaagaaacc gcggcagcta agttcgagag gcagcacatg gacagttaag ctagcttgac
      421 tgactgagat acagcgtacc ttcagctcac agacatgata agatacattg atgagtttgg
      481 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
      541 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca
      601 ttttatgttt caggttcagg gggaggtgtg ggaggttttt taaagcaagt aaaacctcta
      661 caaatgtggt attggcccat ctctatcggt atcgtagcat aaccccttgg ggcctctaaa
      721 cgggtcttga ggggtttttt gtgcccctcg ggccggattg ctatctaccg gcattggcgc
      781 agaaaaaaat gcctgatgcg acgctgcgcg tcttatactc ccacatatgc cagattcagc
      841 aacggatacg gcttccccaa cttgcccact tccatacgtg tcctccttac cagaaattta
      901 tccttaaggt cgtcagctat cctgcaggcg atctctcgat ttcgatcaag acattccttt
      961 aatggtcttt tctggacacc actaggggtc agaagtagtt catcaaactt tcttccctcc
     1021 ctaatctcat tggttacctt gggctatcga aacttaatta accagtcaag tcagctactt
     1081 ggcgagatcg acttgtctgg gtttcgacta cgctcagaat tgcgtcagtc aagttcgatc
     1141 tggtccttgc tattgcaccc gttctccgat tacgagtttc atttaaatca tgtgagcaaa
     1201 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
     1261 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
     1321 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
     1381 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
     1441 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
     1501 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
     1561 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
     1621 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
     1681 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
     1741 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
     1801 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
     1861 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
     1921 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
     1981 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
     2041 agcgatctgt ctatttcgtt catccatagt tgcatttaaa tttccgaact ctccaaggcc
     2101 ctcgtcggaa aatcttcaaa cctttcgtcc gatccatctt gcaggctacc tctcgaacga
     2161 actatcgcaa gtctcttggc cggccttgcg ccttggctat tgcttggcag cgcctatcgc
     2221 caggtattac tccaatcccg aatatccgag atcgggatca cccgagagaa gttcaaccta
     2281 catcctcaat cccgatctat ccgagatccg aggaatatcg aaatcggggc gcgcctggcc
     2341 tccgcgccgg gttttggcgc ctcccgcggg cgcccccctc gtcacggcga gcgctgccac
     2401 gtcagacgaa gggcgcagga gcgtcctgat ccttccgccc ggacgctcag gacagcggcc
     2461 cgctgctcat aagactcggc cttagaaccc cagtatcagc agaaggacat tttaggacgg
     2521 gacttgggtg actctagggc actggttttc tttccagaga gcggaacagg cgaggaaaag
     2581 tagtcccttc tcggcgattc tgcggaggga tctccgtggg gcggtgaacg ccgatgatta
     2641 tataaggacg cgccgggtgt ggcacagcta gttccgtcgc agccgggatt tgggtcgcgg
     2701 ttcttgtttg tggatcgctg tgatcgtcac ttggtgagta gcgggctgct gggctggccg
     2761 gggctttcgt ggccgccggg ccgctcggtg ggacggaagc gtgtggagag accgccaagg
     2821 gctgtagtct gggtccgcga gcaaggttgc cctgaactgg gggttggggg gagcgcagca
     2881 aaatggcggc tgttcccgag tcttgaatgg aagacgcttg tgaggcgggc tgtgaggtcg
     2941 ttgaaacaag gtggggggca tggtgggcgg caagaaccca aggtcttgag cccttcgcta
     3001 atgcgggaaa gctcttattc gggtgagatg ggctgggcac catctgggga ccctgacgtg
     3061 aagtttgtca ctgactggag aactcggttt gtcgtctgtt gcgggggcgg cagttatggc
     3121 ggtgccgttg ggcagtgcac ccgtaccttt gggagcgcgc gccctcgtcg tgtcgtgacg
     3181 tcacccgttc tgttggctta taatgcaggg tggggccacc tgccggtagg tgtgcggtag
     3241 gcttttctcc gtcgcaggac gcagggttcg ggcctagggt aggctctcct gaatcgacag
     3301 gcgccggacc tctggtgagg ggagggataa gtgaggcgtc agtttctttg gtcggtttta
     3361 tgtacctatc ttcttaagta gctgaagctc cggttttgaa ctatgcgctc ggggttggcg
     3421 agtgtgtttt gtgaagtttt ttaggcacct tttgaaatgt aatcatttgg gtcaatatgt
     3481 aattttcagt gttagacttg taaattgtcc gctaaattct ggccgttttt ggcttttttg
     3541 ttagacaaca tgaccgagta caagcccacg gtgcgcctcg ccacccgcga cgacgtcccc
     3601 agggccgtac gcaccctcgc cgccgcgttc gccgactacc ccgccacgcg ccacaccgtc
     3661 gatccggacc gccacatcga gcgggtcacc gagctgcaag aactcttcct cacgcgcgtc
     3721 gggctcgaca tcggcaaggt gtgggtcgcg gacgacggcg ccgcggtggc ggtctggacc
     3781 acgccggaga gcgtcgaagc gggggcggtg ttcgccgaga tcggcccgcg catggccgag
     3841 ttgagcggtt cccggctggc cgcgcagcaa cagatggagg gcctcctggc gccgcaccgg
     3901 cccaaggagc ccgcgtggtt cctggccacc gtcggcgtct cgcccgacca ccagggcaag
     3961 ggtctgggca gcgccgtcgt gctccccgga gtggaggccg ccgagcgcgc cggggtgccc
     4021 gccttcctgg agacctccgc gccccgcaac ctccccttct acgagcggct cggcttcacc
     4081 gtcaccgccg acgtcgaggt gcccgaagga ccgcgcacct ggtgcatgac ccgcaagccc
     4141 ggtgcctagg ctagtatgta agcctagtct tagataataa aatcgctatc catcgaagat
     4201 ggatgtgtgt tggttttttg tgtgtgtaac gctaggcgcg cctggtgtac cgagaacgat
     4261 cctctcagtg cgagtctcga cgatccatat cgttgcttgg cagtcagcca gtcggaatcc
     4321 agcttgggac ccaggaagtc caatcgtcag atattgtact caagcctggt cacggcagcg
     4381 taccgatctg tttaaaccta gatattgata gtctgatcgg tcaacgtata atcgagtcct
     4441 agcttttgca aacatctatc aagagacagg atcagcagga ggctttcgca tgagtattca
     4501 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     4561 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcgc gagtgggtta
     4621 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgctt
     4681 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     4741 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtattc
     4801 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     4861 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgattg gaggaccgaa
     4921 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     4981 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     5041 ggcaacaacc ttgcgtaaac tattaactgg cgaactactt actctagctt cccggcaaca
     5101 gttgatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     5161 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     5221 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     5281 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     5341 gcattggtaa ccgattctag gtgcattggc gcagaaaaaa atgcctgatg cgacgctgcg
     5401 cgtcttatac tcccacatat gccagattca gcaacggata cggcttcccc aacttgccca
     5461 cttccatacg tgtcctcctt accagaaatt tatccttaag atcgtttaaa ctcgactctg
     5521 gctctatcga atctccgtcg tttcgagctt acgcgaacag ccgtggcgct catttgctcg
     5581 tcgggcatcg aatctcgtca gctatcgtca gcttaccttt ttggcagcga tcgcggctcc
     5641 cgacatcttg gaccattagc tccacaggta tcttcttccc tctagtggtc ataacagcag
     5701 cttcagctac ctctcaattc aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt
     5761 atgctatcaa tcgttgcgtt acacacacaa aaaaccaaca cacatccatc ttcgatggat
     5821 agcgatttta ttatctaact gctgatcgag tgtagccaga tctagtaatc aattacgggg
     5881 tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt aaatggcccg
     5941 cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta tgttcccata
     6001 gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg gtaaactgcc
     6061 cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga cgtcaatgac
     6121 ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt tcctacttgg
     6181 cagtacatct acgtattagt catcgctatt accatgctga tgcggttttg gcagtacatc
     6241 aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc cattgacgtc
     6301 aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg taacaactcc
     6361 gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat aagcagagct
     6421 ggtttagtga accgtcagat cagatctttg tcgatcctac catccactcg acacacccgc
     6481 cagcggccgc
//
