LOCUS       sequence-384705-        4641 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses human codon-optimized SpCas9-NRCH and blasticidin 
            resistance: EFS promoter-SpCas9-NRCH-NLS-FLAG-P2A-BSD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 4641)
  AUTHORS   Kim N, Choi S, Kim S, Song M, Seo JH, Min S, Park J, Cho SR, Kim HH
  TITLE     Deep learning models to predict the editing efficiencies and 
            outcomes of diverse base editors.
  JOURNAL   Nat Biotechnol. 2024 Mar;42(3):484-497. doi: 
            10.1038/s41587-023-01792-x
  PUBMED    37188916
REFERENCE   2  (bases 1 to 4641)
  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"; doi: "10"; journalName: 
            "Nat Biotechnol"; date: "2024-03"; volume: "42"; issue: "3"; pages: 
            "484-497"
COMMENT     Sequence Label: SpCas9-NRCH
FEATURES             Location/Qualifiers
     source          1..4641
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             1..4104
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMVKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGIIPHQIHLGELHAILRRQGDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRLRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGG
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKGNSDKLIARKKDWDPKKYGGFNSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGVLQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTINRKQYNTTKEVL
                     DATLIRQSITGLYETRIDLSQLGGD"
     CDS             4105..4152
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4153..4176
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4186..4242
                     /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"
     CDS             4243..4641
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
ORIGIN
        1 atggacaaga agtacagcat cggcctggac atcggcacca actctgtggg ctgggccgtg
       61 atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa caccgaccgg
      121 cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga aacagccgag
      181 gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa ccggatctgc
      241 tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt cttccacaga
      301 ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc catcttcggc
      361 aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca cctgagaaag
      421 aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc cctggcccac
      481 atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga caacagcgac
      541 gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga ggaaaacccc
      601 atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag caagagcaga
      661 cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct gttcggaaac
      721 ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga cctggccgag
      781 gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa cctgctggcc
      841 cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc cgacgccatc
      901 ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct gagcgcctct
      961 atggtgaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg
     1021 cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa cggctacgcc
     1081 ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa gcccatcctg
     1141 gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg
     1201 aagcagcgga ccttcgacaa cggcatcatc ccccaccaga tccacctggg agagctgcac
     1261 gccattctgc ggcggcaggg cgatttttac ccattcctga aggacaaccg ggaaaagatc
     1321 gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc
     1381 agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa
     1441 gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag
     1501 aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg
     1561 tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg
     1621 agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc
     1681 gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc
     1741 tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt
     1801 atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg
     1861 ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc
     1921 cacctgttcg acgacaaagt gatgaagcag ctgaagcggc tgagatacac cggctggggc
     1981 aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg
     2041 gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac
     2101 agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg
     2161 cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca
     2221 gtgaaggtgg tggacgagct cgtgaaagtg atgggcggcc acaagcccga gaacatcgtg
     2281 atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga
     2341 atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc
     2401 gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg
     2461 gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga tgtggaccat
     2521 atcgtgcctc agagctttct gaaggacgac tccatcgaca acaaggtgct gaccagaagc
     2581 gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag
     2641 aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg
     2701 accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag
     2761 ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac
     2821 actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc
     2881 aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac
     2941 taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag
     3001 taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag
     3061 atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc
     3121 aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat ccggaagcgg
     3181 cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt
     3241 gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg
     3301 cagacaggcg gcttcagcaa agagtctatc ctgcccaagg gcaacagcga taagctgatc
     3361 gccagaaaga aggactggga ccctaagaag tacggcggct tcaacagccc caccgtggcc
     3421 tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact gaagagtgtg
     3481 aaagagctac tggggatcac catcatggaa agaagcagct tcgagaagaa tcccatcgac
     3541 tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag
     3601 tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc cggcgtgctg
     3661 cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta cctggccagc
     3721 cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct gtttgtggaa
     3781 cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc caagagagtg
     3841 atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca ccgggataag
     3901 cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa tctgggagcc
     3961 cctgccgcct tcaagtactt tgacaccacc atcaaccgga agcagtacaa caccaccaaa
     4021 gaggtgctgg acgccaccct gatccggcag agcatcaccg gcctgtacga gacacggatc
     4081 gacctgtctc agctgggagg cgacaagcga cctgccgcca caaagaaggc tggacaggct
     4141 aagaagaaga aagattacaa agacgatgac gataagggat ccggcgcaac aaacttctct
     4201 ctgctgaaac aagccggaga tgtcgaagag aatcctggac cgatggccaa gcctttgtct
     4261 caagaagaat ccaccctcat tgaaagagca acggctacaa tcaacagcat ccccatctct
     4321 gaagactaca gcgtcgccag cgcagctctc tctagcgacg gccgcatctt cactggtgtc
     4381 aatgtatatc attttactgg gggaccttgt gcagaactcg tggtgctggg cactgctgct
     4441 gctgcggcag ctggcaacct gacttgtatc gtcgcgatcg gaaatgagaa caggggcatc
     4501 ttgagcccct gcggacggtg ccgacaggtg cttctcgatc tgcatcctgg gatcaaagcc
     4561 atagtgaagg acagtgatgg acagccgacg gcagttggga ttcgtgaatt gctgccctct
     4621 ggttatgtgt gggagggcta a
//
