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Generation of targeted mouse mutants by embryo microinjection of TALEN mRNA

Wefers, Benedikt, Panda, Sudeepta K, Ortiz, Oskar, Brandl, Christina, Hensler, Svenja, Hansen, Jens, Wurst, Wolfgang, Kühn, Ralf·2013·Nature Protocols
DOI 10.1038/nprot.2013.142
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Abstract

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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Metadata

Authors
Wefers, B., Panda, S. K., Ortiz, O., Brandl, C., Hensler, S., Hansen, J., Wurst, W., & Kühn, R.
Venue
Nature Protocols
Publisher
Springer Science and Business Media LLC
Published
2013-10-31
Volume
8
Issue
12
Pages
2355-2379
ISSN
  • 1754-2189
  • 1750-2799
DOI
10.1038/nprot.2013.142
Cited by
58
References
4
Language
en
Licence
www.springer.com/tdm (tdm)

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