LOCUS       sequence-261038-        5352 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expression of nSpyCas9 with N-terminal miniABEMax (V82G).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5352)
  AUTHORS   Nguyen Tran MT, Mohd Khalid MKN, Wang Q, Walker JKR, Lidgerwood GE, 
            Dilworth KL, Lisowski L, Pebay A, Hewitt AW
  TITLE     Engineering domain-inlaid SaCas9 adenine base editors with reduced 
            RNA off-targets and increased on-target DNA editing.
  JOURNAL   Nat Commun. 2020 Sep 25;11(1):4871. doi: 10.1038/s41467-020-18715-y.
  PUBMED    32978399
REFERENCE   2  (bases 1 to 5352)
  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"; journalName: "Nat 
            Commun."; date: "2020-09-25"; volume: "11(1):4871. doi"; pages: " 
            10.1038/s41467-020-18715-y"
COMMENT     Sequence Label: miniABEMax V82G wt
FEATURES             Location/Qualifiers
     source          1..5352
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             49..69
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(664..687,688..690,691..3177,3178..3180,3181..4764)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             4777..4797
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4810..4857
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4858..4881
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4891..4947
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
ORIGIN
        1 aagcttgcca ccatgaaacg gacagccgac ggaagcgagt tcgagtcacc aaagaagaag
       61 cggaaagtct ctgaggtgga gttttcccac gagtactgga tgagacatgc cctgaccctg
      121 gccaagaggg cacgcgatga gagggaggtg cctgtgggag ccgtgctggt gctgaacaat
      181 agagtgatcg gcgagggctg gaacagagcc atcggcctgc acgacccaac agcccatgcc
      241 gaaattatgg ccctgagaca gggcggcctg gtcatgcaga actacagact gattgacgcc
      301 accctgtacg gcacattcga gccttgcgtg atgtgcgccg gcgccatgat ccactctagg
      361 atcggccgcg tggtgtttgg cgtgaggaac gcaaaaaccg gcgccgcagg ctccctgatg
      421 gacgtgctgc actaccccgg catgaatcac cgcgtcgaaa ttaccgaggg aatcctggca
      481 gatgaatgtg ccgccctgct gtgctatttc tttcggatgc ctagacaggt gttcaatgct
      541 cagaagaagg cccagagctc caccgactcc ggaggatcta gcggaggctc ctctggctct
      601 gagacacctg gcacaagcga gagcgcaaca cctgaaagca gcgggggcag cagcgggggg
      661 tcagataaaa agtatagcat tggtctggcg attggcacca acagcgtggg ctgggccgtg
      721 attaccgatg agtacaaagt tccgtccaag aagttcaagg tgctgggtaa caccgaccgc
      781 cacagcatca agaaaaacct gattggtgcc ctgctgtttg atagcggtga aaccgccgag
      841 gctacccgtc tgaagcgcac cgcccgtcgc cgttataccc gtcgtaaaaa ccgtatctgc
      901 tacctgcaag agattttcag caacgaaatg gccaaagtgg acgatagctt ctttcaccgc
      961 ctggaggaga gcttcctggt ggaggaagac aagaaacacg agcgtcaccc gatctttggc
     1021 aacattgtgg atgaggtggc ttatcacgaa aagtacccga ccatctatca cctgcgtaag
     1081 aaactggtgg atagcaccga caaggccgat ctgcgcctga tctacctggc gctggctcac
     1141 atgatcaagt tccgtggtca ctttctgatc gagggcgacc tgaacccgga taacagcgat
     1201 gtggacaagc tgttcatcca gctggtgcag acctataacc agctgtttga ggaaaacccg
     1261 attaacgcca gcggtgtgga cgccaaggcg atcctgagcg cgcgtctgag caaaagccgc
     1321 cgtctggaga acctgatcgc gcagctgccg ggtgaaaaga aaaacggtct gtttggcaac
     1381 ctgattgctc tgagcctggg cctgaccccg aacttcaaaa gcaactttga cctggccgag
     1441 gatgcgaagc tgcaactgag caaggatacc tacgacgatg acctggacaa cctgctggcg
     1501 cagatcggcg accagtatgc tgatctgttc ctggccgcga agaacctgag cgatgcgatc
     1561 ctgctgagcg acattctgcg tgtgaacacc gagattacca aggctccgct gagcgccagc
     1621 atgatcaaac gctacgatga acaccaccag gacctgaccc tgctgaaagc tctggtgcgt
     1681 cagcagctgc cggagaagta taaagaaatc ttctttgacc agagcaagaa cggttacgcc
     1741 ggctatattg atggcggtgc gagccaggag gagttctaca agtttatcaa accgattctg
     1801 gagaaaatgg acggcaccga ggaactgctg gtgaagctga accgtgaaga tctgctgcgt
     1861 aaacagcgca cctttgacaa cggtagcatc ccgcaccaga ttcacctggg tgagctgcac
     1921 gccatcctgc gccgtcagga agatttctac ccgtttctga aggacaaccg tgagaagatc
     1981 gaaaaaattc tgaccttccg cattccgtac tatgtgggtc cgctggctcg tggcaacagc
     2041 cgctttgcct ggatgacccg caagagcgag gaaaccatca ccccgtggaa cttcgaggaa
     2101 gtggtggaca aaggcgccag cgcccagagc ttcattgagc gtatgaccaa ctttgataag
     2161 aacctgccga acgaaaaagt gctgccgaag cacagcctgc tgtacgagta tttcaccgtg
     2221 tataacgaac tgaccaaggt gaaatacgtg accgagggta tgcgcaagcc ggccttcctg
     2281 agcggcgaac agaagaaagc gatcgtggac ctgctgttta aaaccaaccg taaggtgacc
     2341 gtgaagcagc tgaaagagga ttacttcaag aaaattgaat gcttcgacag cgtggagatc
     2401 agcggtgtgg aagatcgttt taacgcgagc ctgggcacct atcacgacct gctgaaaatc
     2461 attaaggata aagacttcct ggataacgag gaaaacgagg acatcctgga agatattgtg
     2521 ctgaccctga ccctgtttga ggaccgtgaa atgatcgagg aacgcctgaa gacctacgcc
     2581 cacctgttcg atgacaaagt gatgaaacag ctgaagcgcc gtcgctatac cggttggggc
     2641 cgtctgagcc gcaaactgat caacggtatt cgcgataaac agagcggcaa gaccatcctg
     2701 gattttctga agagcgacgg tttcgcgaac cgtaacttta tgcagctgat tcacgatgac
     2761 agcctgacct tcaaagagga catccagaag gcgcaggtga gcggtcaggg cgatagcctg
     2821 cacgaacaca tcgctaacct ggccggtagc ccggccatca agaagggcat cctgcaaacc
     2881 gtgaaggtgg tggacgagct ggtgaaagtg atgggtcgtc acaagccgga aaacatcgtg
     2941 attgagatgg cccgcgaaaa ccagaccacc cagaaaggcc agaagaacag ccgtgagcgc
     3001 atgaagcgta tcgaggaagg catcaaggaa ctgggcagcc agatcctgaa ggagcacccg
     3061 gtggaaaaca cccagctgca aaacgagaaa ctgtatctgt actacctgca aaacggtcgt
     3121 gacatgtacg tggatcagga gctggacatt aaccgcctga gcgattacga cgtggatgcg
     3181 atcgtgccgc agagcttcct gaaggatgac agcattgaca acaaagtgct gacccgtagc
     3241 gataaaaacc gcggcaagag cgacaacgtg ccgagcgagg aagtggtgaa gaaaatgaaa
     3301 aactactggc gtcagctgct gaacgccaaa ctgatcaccc agcgtaagtt tgacaacctg
     3361 accaaagctg agcgcggcgg tctgagcgaa ctggataaag ccggtttcat taagcgtcag
     3421 ctggtggaaa cccgccagat caccaagcac gtggcgcaga ttctggacag ccgcatgaac
     3481 accaagtacg acgagaacga taaactgatc cgtgaagtga aagtgatcac cctgaagagc
     3541 aaactggtga gcgattttcg taaagacttc cagttttaca aggtgcgcga gatcaacaac
     3601 tatcaccacg cgcacgacgc ttacctgaac gccgtggtgg gcaccgccct gatcaagaag
     3661 tatccgaagc tggagagcga gttcgtgtac ggcgattata aagtgtacga cgtgcgtaaa
     3721 atgatcgcga agagcgagca ggaaattggc aaagcgaccg ctaagtattt cttttacagc
     3781 aacatcatga acttctttaa gaccgagatc accctggcca acggtgaaat ccgtaaacgc
     3841 ccgctgattg aaaccaacgg tgaaaccggc gaaatcgtgt gggataaggg ccgtgacttt
     3901 gcgaccgtgc gcaaagtgct gtccatgccg caggtgaaca ttgtgaagaa aaccgaggtg
     3961 cagaccggcg gtttcagcaa ggaaagcatt ctgccgaaac gtaacagcga caagctgatc
     4021 gcccgcaaga aagactggga cccgaagaag tatggcggtt tcgatagccc aaccgtggcc
     4081 tacagcgtgc tggtggtggc taaagtggag aagggtaaaa gcaagaaact gaaaagcgtg
     4141 aaggaactgc tgggcatcac cattatggag cgtagcagct tcgaaaagaa cccgatcgac
     4201 tttctggagg cgaaaggtta caaggaagtg aagaaagatc tgatcatcaa gctgccgaag
     4261 tatagcctgt ttgagctgga aaatggtcgt aaacgtatgc tggccagcgc cggcgagctg
     4321 caaaagggca acgaactggc gctgccgagc aaatacgtga acttcctgta tctggccagc
     4381 cactacgaga agctgaaagg tagcccggag gacaacgaac agaagcagct gtttgtggag
     4441 cagcacaaac actacctgga tgagatcatt gaacagatta gcgagttcag caagcgtgtg
     4501 atcctggctg atgccaacct ggacaaagtg ctgagcgcct ataacaaaca ccgtgacaag
     4561 ccgatccgcg agcaggcgga aaacatcatt cacctgttca ccctgaccaa cctgggtgcg
     4621 ccggctgcct tcaagtattt tgataccacc attgaccgta aacgctacac cagcaccaag
     4681 gaggtgctgg atgctaccct gatccaccag agcattaccg gcctgtacga aacccgtatc
     4741 gatctgagcc agctgggcgg tgactctggt ggttctccca agaagaagag gaaagtctct
     4801 ggtggttcta agcgtccggc ggctaccaag aaagccggtc aggcgaagaa aaagaaagac
     4861 tacaaagatg acgatgacaa gggtagcggc gctaccaact tcagcctgct gaagcaggcg
     4921 ggcgacgtgg aggaaaatcc gggcccgatg gctaaaccgc tgagccagga ggaaagcacc
     4981 ctgatcgagc gcgcgaccgc taccattaac agcatcccga ttagcgaaga ttatagcgtg
     5041 gccagcgccg cgctgagcag cgatggtcgt atcttcaccg gcgtgaacgt gtatcacttt
     5101 accggtggcc cgtgcgccga gctggtggtg ctgggcaccg ccgccgccgc cgccgccggc
     5161 aacctgacct gcatcgtggc gattggtaac gaaaatcgtg gcatcctgag cccgtgcggc
     5221 cgttgccgcc aggtgctgct ggacctgcat ccgggcatca aggcgattgt gaaggatagc
     5281 gacggccaac cgaccgctgt gggtattcgt gaactgctgc catccggcta tgtttgggaa
     5341 ggctaaggta cc
//
