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Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.
Generation of targeted mouse mutants by embryo microinjection of TALEN mRNA.
Nat. Protoc. 8, 2355-2379 (2013)
Genetically engineered mice are instrumental for the analysis of mammalian gene function in health and disease. As classical gene targeting, which is performed in embryonic stem (ES) cell cultures and generates chimeric mice, is a time-consuming and labor-intensive procedure, we recently used transcription activator-like (TAL) effector nucleases (TALENs) for mutagenesis of the mouse genome directly in one-cell embryos. Here we describe a stepwise protocol for the generation of knock-in and knockout mice, including the selection of TALEN-binding sites, the design and construction of TALEN coding regions and of mutagenic oligodeoxynucleotides (ODNs) and targeting vectors, mRNA production, embryo microinjection and the identification of modified alleles in founder mutants and their progeny. After a setup time of 2-3 weeks of hands-on work for TALEN construction, investigators can obtain first founder mutants for genes of choice within 7 weeks after embryo microinjections.
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Publikationstyp
Artikel: Journalartikel
Dokumenttyp
Wissenschaftlicher Artikel
Schlagwörter
Zinc-finger Nucleases ; Homologous Recombination ; Effector Nucleases ; Mammalian-cells ; Iii Effectors ; Dna ; Genome ; Mice ; Mutations ; Frequency
ISSN (print) / ISBN
1754-2189
e-ISSN
1750-2799
Zeitschrift
Nature Protocols
Quellenangaben
Band: 8,
Heft: 12,
Seiten: 2355-2379
Verlag
Nature Publishing Group
Nichtpatentliteratur
Publikationen
Begutachtungsstatus
Peer reviewed
Institut(e)
Institute of Developmental Genetics (IDG)