title: Gene targeting by homologous recombination in mouse zygotes mediated by zinc-finger nucleases.
abstract: Gene targeting by homologous recombination in embryonic stem cells is extensively used to generate specific mouse mutants. However, most mammalian species lack tools for targeted gene manipulation. Since double-strand breaks strongly increase the rate of homologous recombination at genomic loci, we explored whether gene targeting can be directly performed in zygotes by the use of zinc-finger nucleases. Here we report that gene targeting is achieved in 1.7-4.5% of murine one-cell embryos upon the coinjection of targeting vectors with zinc-finger nucleases, without preselection. These findings enable the manipulation of the mammalian germ line in a single step in zygotes, independent of ES cells.
authors.affiliation: Institute for Developmental Genetics, Helmholtz Center Munich, Munich, Germany.
authors.forename: Melanie
authors.initials: M
authors.lastname: Meyer
authors.forename: Martin Hrabé
authors.initials: MH
authors.lastname: de Angelis
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Wurst
authors.forename: Ralf
authors.initials: R
authors.lastname: Kühn
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D004247:DNA / Q000235:genetics; D053595:Embryonic Stem Cells / Q000378:metabolism; D004720:Endonucleases / Q000378:metabolism*; D005260:Female; D018390:Gene Targeting / Q000379:methods*; D017930:Genes, Reporter; D005822:Genetic Vectors; D008164:Luminescent Proteins / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D011247:Pregnancy; D011506:Proteins / Q000235:genetics; D022661:RNA, Untranslated; D011995:Recombination, Genetic; D016335:Zinc Fingers; D015053:Zygote / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; C110853:Gt(ROSA)26Sor non-coding RNA, mouse; D008164:Luminescent Proteins; D011506:Proteins; D022661:RNA, Untranslated; C054562:yellow fluorescent protein, Bacteria; D004247:DNA; D004720:Endonucleases