Targeted genome modification of crop plants using a CRISPR-Cas system

Journal Article / 2013

Shan, Qiwei; Wang, Yanpeng; Li, Jun; Zhang, Yi; Chen, Kunling; Liang, Zhen; Zhang, Kang; Liu, Jinxing; Xi, Jianzhong Jeff; Qiu, Jin-Long; Gao, Caixia

Although genome editing technologies using zinc finger nucleases (ZFNs)1 and transcription activator-like effector nucleases (TALENs)2 can generate genome modifications, new technologies that are robust, affordable and easy to engineer are needed. Recent advances in the study of the prokaryotic adaptive immune system, involving type II clustered, regularly interspaced, short palindromic repeats (CRISPR), provide an alternative genome editing strategy3. Type II CRISPR systems are widespread in bacteria; they use a single endonuclease, a CRISPR-associated protein Cas9, to provide a defense against invading viral and plasmid DNAs4. Cas9 can form a complex with a synthetic single-guide RNA (sgRNA), consisting of a fusion of CRISPR RNA (crRNA) and trans-activating crRNA. The sgRNA guides Cas9 to recognize and cleave target DNA. Cas9 has a HNH nuclease domain and a RuvC-like domain; each cleaves one strand of a doublestranded DNA. It can be used as an RNAguided endonuclease to perform sequencespecific genome editing in bacteria, human cells, zebrafish and mice5–11. Here we 686 show that customizable sgRNAs can direct Cas9 to induce sequence-specific genome modifications in the two most widely cultivated food crops, rice (Oryza sativa) and common wheat (Triticum aestivum).