title: Highly efficient gene knockout in mice and zebrafish with RNA-guided endonucleases.
abstract: RNA-guided endonucleases (RGENs), derived from the prokaryotic Type II CRISPR-Cas system, enable targeted genome modification in cells and organisms. Here we describe the establishment of gene-knockout mice and zebrafish by the injection of RGENs as Cas9 protein:guide RNA complexes or Cas9 mRNA plus guide RNA into one-cell-stage embryos of both species. RGENs efficiently generated germline transmittable mutations in up to 93% of newborn mice with minimal toxicity. RGEN-induced mutations in the mouse Prkdc gene that encodes an enzyme critical for DNA double-strand break repair resulted in immunodeficiency both in F₀ and F₁ mice. We propose that RGEN-mediated mutagenesis in animals will greatly expedite the creation of genetically engineered model organisms, accelerating functional genomic research.
authors.affiliation: Department of Biochemistry, College of Life Science and Biotechnology, Yonsei University, Seoul 120-749, Republic of Korea;
authors.forename: Young Hoon
authors.initials: YH
authors.lastname: Sung
authors.forename: Jong Min
authors.initials: JM
authors.lastname: Kim
authors.forename: Hyun-Taek
authors.initials: HT
authors.lastname: Kim
authors.forename: Jaehoon
authors.initials: J
authors.lastname: Lee
authors.forename: Jisun
authors.initials: J
authors.lastname: Jeon
authors.forename: Young
authors.initials: Y
authors.lastname: Jin
authors.forename: Jung-Hwa
authors.initials: JH
authors.lastname: Choi
authors.forename: Young Ho
authors.initials: YH
authors.lastname: Ban
authors.forename: Sang-Jun
authors.initials: SJ
authors.lastname: Ha
authors.forename: Cheol-Hee
authors.initials: CH
authors.lastname: Kim
authors.forename: Han-Woong
authors.initials: HW
authors.lastname: Lee
authors.forename: Jin-Soo
authors.initials: JS
authors.lastname: Kim
mesh_terms: D000818:Animals; D000831:Animals, Newborn / Q000235:genetics; D064130:CRISPR-Associated Proteins / Q000378:metabolism*; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D004622:Embryo, Mammalian; D004625:Embryo, Nonmammalian; D004720:Endonucleases / Q000235:genetics* / Q000378:metabolism; D051858:Forkhead Transcription Factors / Q000378:metabolism; D055786:Gene Knockout Techniques; D016678:Genome; D018095:Germ-Line Mutation; D051379:Mice; D008807:Mice, Inbred BALB C; D018345:Mice, Knockout; D016296:Mutagenesis; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D010641:Phenotype; D015027:Zebrafish / Q000235:genetics / Q000378:metabolism; D058727:RNA, Small Untranslated
chemical_list: D064130:CRISPR-Associated Proteins; D051858:Forkhead Transcription Factors; D009687:Nuclear Proteins; D004720:Endonucleases; D058727:RNA, Small Untranslated