LOCUS       sequence-62935-e         980 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  This plasmid separately encodes a human codon-optimized SpCas9 
            nickase, a tracrRNA and customizable crRNA..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 980)
  AUTHORS   Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, Hsu PD, Wu X, 
            Jiang W, Marraffini LA, Zhang F
  TITLE     Multiplex Genome Engineering Using CRISPR/Cas Systems.
  JOURNAL   Science. 2013 Jan 3.
  PUBMED    23287718
REFERENCE   2  (bases 1 to 980)
  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: "Science. 
            2013 Jan 3."
COMMENT     Sequence Label: 
            pX334-U6-DR-BB-DR-Cbh-NLS-hSpCas9n(D10A)-NLS-H1-shorttracr-PGK-puro
FEATURES             Location/Qualifiers
     source          1..980
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        complement(26..525)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(74..93)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     467..490
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     protein_bind    complement(586..619)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(695..716)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_RNA        complement(790..868)
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     promoter        complement(879..969)
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     complement(939..958)
                     /label=H1
                     /note="Human H1 promoter, forward primer"
ORIGIN
        1 gctcgagatc tagatggatg caggtcgaaa ggcccggaga tgaggaagag gagaacagcg
       61 cggcagacgt gcgcttttga agcgtgcaga atgccgggcc tccggaggac cttcgggcgc
      121 ccgccccgcc cctgagcccg cccctgagcc cgcccccgga cccacccctt cccagcctct
      181 gagcccagaa agcgaaggag caaagctgct attggccgct gccccaaagg cctacccgct
      241 tccattgctc agcggtgctg tccatctgca cgagactagt gagacgtgct acttccattt
      301 gtcacgtcct gcacgacgcg agctgcgggg cgggggggaa cttcctgact aggggaggag
      361 tagaaggtgg cgcgaagggg ccaccaaaga acggagccgg ttggcgccta ccggtggatg
      421 tggaatgtgt gcgaggccag aggccacttg tgtagcgcca agtgcccagc ggggctgcta
      481 aagcgcatgc tccagactgc cttgggaaaa gcgcctcccc tacccggtag aatttcgagg
      541 tcgagatcct aagcttggct ggacgtaaac tcctcttcag acctaataac ttcgtatagc
      601 atacattata cgaagttata ttaagggtta ttgaatatga tcggaattct tacttgtaca
      661 gctcgtccat gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat
      721 cgcgcttctc gttggggtct ttgctcaggg cggactgggt gctcaggtag tggttgtcgg
      781 cgccaaaaag caccgactcg gtgccacttt ttcaagttga taacggacta gccttatttt
      841 aacttgctat gctgttttga atggttccaa ctagatctgt ggtctcatac agaacttata
      901 agattcccaa atccaaagac atttcacgtt tatggtgatt tcccagaaca catagcgaca
      961 tgcaaatatt gcagggcgcc
//
