Targeted Genome Editing in Crop Plants Using a CRISPR/Cas System

Although genome editing technologies using ZFNs (zinc finger nucleases) and TALENs (transcription activator-like effector nucleases) can generate genome modifications, new technologies that are robust, affordable and easy to engineer are needed. Recent advances in the study adaptive immune system involving type II prokaryotic clustered regularly interspaced short palindromic repeats (CRISPR) provide an alternative genome editing strategy. We describe the use of the CRISPR/Cas system involving RNA-guided endonucleases to introduce specific mutations in rice and wheat. For four rice genes and one wheat gene, the frequencies of sgRNA:Cas9 induced mutations were 14.5 to 38.0% in protoplasts, and 4.0 to 9.4% in regenerated rice plants. We also demonstrate homology-directed targeted gene modification in rice using a single-stranded DNA (ssDNA) oligo as a donor template. Our findings establish that the sgRNA:Cas9 system can be used for rice and wheat genome modification, the first plants shown to be amenable to this gene editing approach.