LOCUS       sequence-70360-e        1033 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  This plasmid separately encodes a human codon-optimized SpCas9, a 
            tracrRNA and customizable crRNA..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 1033)
  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 1033)
  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: 
            pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro
FEATURES             Location/Qualifiers
     source          1..1033
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     5..28
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     protein_bind    complement(124..157)
                     /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(233..254)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_RNA        complement(328..406)
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     promoter        complement(417..631)
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     complement(477..496)
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     polyA_signal    645..852
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     841..858
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(886..933)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
ORIGIN
        1 tgcccagcgg ggctgctaaa gcgcatgctc cagactgcct tgggaaaagc gcctccccta
       61 cccggtagaa tttcgaggtc gagatcctaa gcttggctgg acgtaaactc ctcttcagac
      121 ctaataactt cgtatagcat acattatacg aagttatatt aagggttatt gaatatgatc
      181 ggaattctta cttgtacagc tcgtccatgc cgagagtgat cccggcggcg gtcacgaact
      241 ccagcaggac catgtgatcg cgcttctcgt tggggtcttt gctcagggcg gactgggtgc
      301 tcaggtagtg gttgtcggcg ccaaaaagca ccgactcggt gccacttttt caagttgata
      361 acggactagc cttattttaa cttgctatgc tgttttgaat ggttccaact agatctgtgg
      421 tctcatacag aacttataag attcccaaat ccaaagacat ttcacgttta tggtgatttc
      481 ccagaacaca tagcgacatg caaatattgc agggcgccac tcccctgtcc ctcacagcca
      541 tcttcctgcc agggcgcacg cgcgctgggt gttcccgcct agtgacactg ggcccgcgat
      601 tccttggagc gggttgatga cgtcagcgtt cgaattgcgg ccgctcccca gcatgcctgc
      661 tattctcttc ccaatcctcc cccttgctgt cctgccccac cccacccccc agaatagaat
      721 gacacctact cagacaatgc gatgcaattt cctcatttta ttaggaaagg acagtgggag
      781 tggcaccttc cagggtcaag gaaggcacgg gggaggggca aacaacagat ggctggcaac
      841 tagaaggcac agtcgaggct gatcagcgag ctctaggaat tcttactttt tcttttttgc
      901 ctggccggcc tttttcgtgg ccgccggcct tttgtcgcct cccagctgag acaggtcgat
      961 ccgtgtctcg tacaggccgg tgatgctctg gtggatcagg gtggcgtcca gcacctcttt
     1021 ggtgctggtg tac
//
