LOCUS       sequence-90732-e        2442 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses truncated N-terminal SpCas9 domain fused to a GyrA intein,
            flanked by ITRs for AAV packaging. Combine with C-Terminal Split 
            Cas9 Gyra Intein for full length SpCas9 production.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 2442)
  AUTHORS   Fine EJ, Appleton CM, White DE, Brown MT, Deshmukh H, Kemp ML, Bao G
  TITLE     Trans-spliced Cas9 allows cleavage of HBB and CCR5 genes in human 
            cells using compact expression cassettes.
  JOURNAL   Sci Rep. 2015 Jul 1;5:10777. doi: 10.1038/srep10777.
  PUBMED    26126518
REFERENCE   2  (bases 1 to 2442)
  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: "Sci Rep."; 
            date: "2015-07-1"; volume: "5:10777. doi"; pages: " 
            10.1038/srep10777"
COMMENT     Sequence Label: N-Terminal Split Cas9 with GyrA intein
FEATURES             Location/Qualifiers
     source          1..2442
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     CDS             4..69
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             76..96
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             121..1836
                     /codon_start=1
                     /product="N-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(N)
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIE"
ORIGIN
        1 atggactata aggaccacga cggagactac aaggatcatg atattgatta caaagacgat
       61 gacgataaga tggccccaaa gaagaagcgg aaggtcggta tccacggagt cccagcagcc
      121 gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc
      181 accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac
      241 agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc
      301 acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat
      361 ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg
      421 gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac
      481 atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa
      541 ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg
      601 atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg
      661 gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc
      721 aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg
      781 ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg
      841 attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat
      901 gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag
      961 atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg
     1021 ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg
     1081 atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag
     1141 cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc
     1201 tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa
     1261 aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag
     1321 cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc
     1381 attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag
     1441 aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga
     1501 ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg
     1561 gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac
     1621 ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat
     1681 aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc
     1741 ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg
     1801 aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc
     1861 ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc
     1921 aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg
     1981 accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac
     2041 ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatactgtat caccggtgat
     2101 gctctggtgg cgctgcccga aggcgaatca gtgcgcatcg ctgacatcgt cccgggcgcc
     2161 cgccctaata gcgataacgc catcgacctg aaggtgcttg accgccacgg caatcctgtt
     2221 ctggccgata gactcttcca cagtggtgaa catcctgtgt ataccgtccg caccgtagaa
     2281 ggcctgagag tgacaggcac agccaaccac ccccttttgt gtctggtgga tgttgccggc
     2341 gtccccaccc tgctgtggaa actgattgat gagataaaac ccggagacta tgcagtgatc
     2401 caaaggtccg catttagtgt ggattgcgct ggcttcgctc gc
//
