Tailor-made mutations in Arabidopsis using zinc finger nucleases.

Methods in molecular biology (Clifton, N.J.) (2013-09-24)
Yiping Qi, Colby G Starker, Feng Zhang, Nicholas J Baltes, Daniel F Voytas

Zinc finger nucleases (ZFNs) are proteins engineered to make site-specific double-strand breaks (DSBs) in a DNA sequence of interest. Imprecise repair of the ZFN-induced DSBs by the nonhomologous end-joining (NHEJ) pathway results in a spectrum of mutations, such as nucleotide substitutions, insertions, and deletions. Here we describe a method for targeted mutagenesis in Arabidopsis with ZFNs, which are engineered by context-dependent assembly (CoDA). This ZFN-induced mutagenesis method is an alternative to other currently available gene knockout or knockdown technologies and is useful for reverse genetic studies.

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Nde I from Neisseria denitrificans, Restriction Enzyme