LOCUS       sequence-80642-e         921 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Expresses human codon-optimized Cas9 protein and puromycin 
            resistance from EFS promoter and an EGFP targeting synthetic 
            single-guide RNA (sgRNA) element from U6 promoter. Lentiviral 
            backbone..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 921)
  AUTHORS   Shalem O, Sanjana NE, Hartenian E, Shi X, Scott DA, Mikkelsen TS, 
            Heckl D, Ebert BL, Root DE, Doench JG, Zhang F
  TITLE     Genome-scale CRISPR-Cas9 knockout screening in human cells.
  JOURNAL   Science. 2014 Jan 3;343(6166):84-7. doi: 10.1126/science.1247005. 
            Epub 2013 Dec 12.
  PUBMED    24336571
REFERENCE   2  (bases 1 to 921)
  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: 
            "10.1126/science.1247005"; journalName: "Science"; date: "2014-01-3-
            3"; volume: "343"; issue: "6166"; pages: "84-7"
COMMENT     Sequence Label: lentiCRISPR - EGFP sgRNA 3
FEATURES             Location/Qualifiers
     source          1..921
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     133..152
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        231..306
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    354..471
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        540..751
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      771..780
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             780..803
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             810..830
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac aaagatatta
       61 gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta
      121 tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct
      181 ttatatatct tgtggaaagg acgaaacacc ggccacaagt tcagcgtgtc gttttagagc
      241 tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt
      301 cggtgctttt ttgaattcta gatcttgaga caaatggcag tattcatcca caattttaaa
      361 agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca
      421 gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat
      481 tacagggaca gcagagatcc actttggcgc cggctcgagt ggctccggtg cccgtcagtg
      541 ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac
      601 cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
      661 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
      721 ttttcgcaac gggtttgccg ccagaacaca ggtgtcgtga cgcgggatcc gccaccatgg
      781 attacaaaga cgatgacgat aagatggccc caaagaagaa gcggaaggtc ggtatccacg
      841 gagtcccagc agccgacaag aagtacagca tcggcctgga catcggcacc aactctgtgg
      901 gctgggccgt gatcaccgac g
//
