LOCUS       sequence-80644-e         998 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 998)
  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 998)
  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 2
FEATURES             Location/Qualifiers
     source          1..998
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     135..154
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        233..308
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    356..473
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        542..753
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      773..782
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             782..805
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             812..832
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 tacgatacaa ggctgttaga gagataatta gaattaattt gactgtaaac acaaagatat
       61 tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat
      121 tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg
      181 ctttatatat cttgtggaaa ggacgaaaca ccgagctgga cggcgacgta aagttttaga
      241 gctagaaata gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga
      301 gtcggtgctt ttttgaattc tagatcttga gacaaatggc agtattcatc cacaatttta
      361 aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa
      421 cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt
      481 attacaggga cagcagagat ccactttggc gccggctcga gtggctccgg tgcccgtcag
      541 tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt cggcaattga
      601 accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt gtactggctc
      661 cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt
      721 ctttttcgca acgggtttgc cgccagaaca caggtgtcgt gacgcgggat ccgccaccat
      781 ggattacaaa gacgatgacg ataagatggc cccaaagaag aagcggaagg tcggtatcca
      841 cggagtccca gcagccgaca agaagtacag catcggcctg gacatcggca ccaactctgt
      901 gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg
      961 caacaccgac cggcacagca tcaagaagaa cctgatcg
//
