title: Efficient gene targeting of the Rosa26 locus in mouse zygotes using TALE nucleases.
abstract: Gene targeting in mice mainly employs homologous recombination (HR) in embryonic stem (ES) cells. Although it is a standard way for production of genetically modified mice, the procedure is laborious and time-consuming. This study describes targeting of the mouse Rosa26 locus by transcription activator-like effector nucleases (TALENs). We employed TALEN-assisted HR in zygotes to introduce constructs encoding TurboRFP and TagBFP fluorescent proteins into the first intron of the Rosa26 gene, and in this way generated two transgenic mice. We also demonstrated that these Rosa26-specific TALENs exhibit high targeting efficiency superior to that of zinc-finger nucleases (ZFNs) specific for the same targeting sequence. Moreover, we devised a reporter assay to assess TALENs activity and specificity to improve the quality of TALEN-assisted targeting.
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic.
authors.forename: Petr
authors.initials: P
authors.lastname: Kasparek
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic.
authors.forename: Michaela
authors.initials: M
authors.lastname: Krausova
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic.
authors.forename: Radka
authors.initials: R
authors.lastname: Haneckova
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic.
authors.forename: Vitezslav
authors.initials: V
authors.lastname: Kriz
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic.
authors.forename: Olga
authors.initials: O
authors.lastname: Zbodakova
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic. Electronic address: vladimir.korinek@img.cas.cz.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Korinek
authors.affiliation: Institute of Molecular Genetics of the AS CR, v.v.i., Videnska 1083, 142 20, Czech Republic. Electronic address: radislav.sedlacek@img.cas.cz.
authors.forename: Radislav
authors.initials: R
authors.lastname: Sedlacek
mesh_terms: D000818:Animals; D001483:Base Sequence; D004720:Endonucleases / Q000378:metabolism*; D018390:Gene Targeting / Q000379:methods*; D056426:Genetic Loci / Q000235:genetics*; D059765:Homologous Recombination; D051379:Mice; D008822:Mice, Transgenic; D015053:Zygote / Q000378:metabolism*
chemical_list: D004720:Endonucleases
keywords: Gene targeting; Homologous recombination; Rosa26 locus; Transcription activator-like effector nucleases; Transgenic; Zinc-finger nucleases