Gene targeting by homologous recombination in mouse zygotes mediated by zinc-finger nucleases.
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.
Meyer
Melanie
M
Institute for Developmental Genetics, Helmholtz Center Munich, Munich, Germany.
de Angelis
Martin Hrabé
MH
Wurst
Wolfgang
W
Kühn
Ralf
R
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*
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