LOCUS       sequence-99323-e         629 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses Cas9 nickase and gRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 629)
  AUTHORS   Sakuma T, Nishikawa A, Kume S, Chayama K, Yamamoto T
  TITLE     Multiplex genome engineering in human cells using all-in-one 
            CRISPR/Cas9 vector system.
  JOURNAL   Sci Rep. 2014 Jun 23;4:5400. doi: 10.1038/srep05400.
  PUBMED    24954249
REFERENCE   2  (bases 1 to 629)
  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: "2014-06-23"; volume: "4:5400. doi"; pages: " 
            10.1038/srep05400"
COMMENT     Sequence Label: pX330A_D10A-1x2
FEATURES             Location/Qualifiers
     source          1..629
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     complement(16..36)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(16..34)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(55..71)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(55..71)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(68..90)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    79..95
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(103..109,110..127,128..133))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    148..169
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     enhancer        223..508
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
ORIGIN
        1 ctccagcttt tgttcccttt agtgagggtt aattgcgcgc ttggcgtaat catggtcata
       61 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
      121 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
      181 ctcactgccc gctttccacc ggtggtctct tctagaggta cccgttacat aacttacggt
      241 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa tagtaacgcc
      301 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      361 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      421 gcccgcctgg cattgtgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      481 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
      541 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      601 tgcagcgatg ggggcggggg ggggggggg
//
