LOCUS       sequence-96722-e         909 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Cas9n (D10A nickase mutant) from S. pyogenes with 2A-EGFP, and 
            cloning backbone for sgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 909)
  AUTHORS   Ran FA, Hsu PD, Wright J, Agarwala V, Scott DA, Zhang F
  TITLE     Genome engineering using the CRISPR-Cas9 system.
  JOURNAL   Nat Protoc. 2013 Nov;8(11):2281-308. doi: 10.1038/nprot.2013.143. 
            Epub 2013 Oct 24.
  PUBMED    24157548
REFERENCE   2  (bases 1 to 909)
  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: 
            "10.1038/nprot.2013.143"; journalName: "Nat Protoc"; date: 
            "2013-11"; volume: "8"; issue: "11"; pages: "2281-308"
COMMENT     Sequence Label: pSpCas9n(BB)-2A-GFP (PX461)
FEATURES             Location/Qualifiers
     source          1..909
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        124..372
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     124..144
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     295..314
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        391..466
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        563..848
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
ORIGIN
        1 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
       61 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
      121 tgtgagggcc tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga
      181 gagataattg gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt
      241 agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat
      301 catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa
      361 ggacgaaaca ccgggtcttc gagaagacct gttttagagc tagaaatagc aagttaaaat
      421 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgttttaga
      481 gctagaaata gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc caattctgca
      541 gacaaatggc tctagaggta cccgttacat aacttacggt aaatggcccg cctggctgac
      601 cgcccaacga cccccgccca ttgacgtcaa tagtaacgcc aatagggact ttccattgac
      661 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      721 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattgtgccc
      781 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      841 ttaccatggt cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc
      901 cacccccaa
//
