LOCUS       sequence-122958-         955 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Cas9 from S. pyogenes with 2A-Puro, and cloning backbone for sgRNA 
            (V2.0).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 955)
  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 955)
  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: pSpCas9(BB)-2A-Puro (PX459) V2.0
FEATURES             Location/Qualifiers
     source          1..955
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     69..86
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(114..713)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(197..217)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     694..713
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     CDS             complement(714..767)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(783..830)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
ORIGIN
        1 agtgggagtg gcaccttcca gggtcaagga aggcacgggg gaggggcaaa caacagatgg
       61 ctggcaacta gaaggcacag tcgaggctga tcagcgagct ctagttagaa ttctcaggca
      121 ccgggcttgc gggtcatgca ccaggtgcgc ggtccttcgg gcacctcgac gtcggcggtg
      181 acggtgaagc cgagccgctc gtagaagggg aggttgcggg gcgcggaggt ctccaggaag
      241 gcgggcaccc cggcgcgctc ggccgcctcc actccgggga gcacgacggc gctgcccaga
      301 cccttgccct ggtggtcggg cgagactccg acggtggcca ggaaccacgc gggctccttg
      361 ggccggtgcg gcgccaggag gccttccatc tgttgctgcg cggccagccg ggaaccgctc
      421 aactcggcca tgcgcgggcc gatctcggcg aacaccgccc ccgcttcgac gctctccggc
      481 gtggtccaga ccgccaccgc ggcgccgtcg tccgcgaccc acaccttgcc gatgtcgagc
      541 ccgacgcgcg tgaggaagag ttcttgcagc tcggtgaccc gctcgatgtg gcggtccgga
      601 tcgacggtgt ggcgcgtggc ggggtagtcg gcgaacgcgg cggcgagggt gcgtacggcc
      661 ctggggacgt cgtcgcgggt ggcgaggcgc accgtgggct tgtactcggt cattgggcca
      721 ggattctcct cgacgtcacc gcatgttagc agacttcctc tgccctctcc actgccgaat
      781 tcctttttct tttttgcctg gccggccttt ttcgtggccg ccggcctttt gtcgcctccc
      841 agctgagaca ggtcgatccg tgtctcgtac aggccggtga tgctctggtg gatcagggtg
      901 gcgtccagca cctctttggt gctggtgtac ctcttccggt cgatggtggt gtcaa
//
