Precise genome modification in the crop species Zea mays using zinc-finger nucleases

Journal Article / 2009

Shukla, Vipula K.; Doyon, Yannick; Miller, Jeffrey C.; DeKelver, Russell C.; Moehle, Erica A.; Worden, Sarah E.; Mitchell, Jon C.; Arnold, Nicole L.; Gopalan, Sunita; Meng, Xiangdong; Choi, Vivian M.; Rock, Jeremy M.; Wu, Ying-Ying; Katibah, George E.; Zhifang, Gao; McCaskill, David; Simpson, Matthew A.; Blakeslee, Beth; Greenwalt, Scott A.; Butler, Holly J.; Hinkley, Sarah J.; Zhang, Lei; Rebar, Edward J.; Gregory, Philip D.; Urnov, Fyodor D.

Agricultural biotechnology is limited by the inefficiencies of conventional random mutagenesis and transgenesis. Because targeted genome modification in plants has been intractable1 , plant trait engineering remains a laborious, time-consuming and unpredictable undertaking. Here we report a broadly applicable, versatile solution to this problem: the use ofdesigned zinc-finger nucleases (ZFNs) that induce a double-stranded break at their target locus2. We describe the use of ZFNs to modify endogenous loci in plants of the crop species Zea mays. We show that simultaneous expression ofZFNs and delivery ofa simple heterologous donor molecule leads to precise targeted addition ofan herbicide-tolerance gene at the intended locus in a significant number ofisolated events. ZFNmodified maize plants faithfully transmit these genetic changes to the next generation. Insertional disruption of one target locus, IPK1, results in both herbicide tolerance and the expected alteration of the inositol phosphate profile in developing seeds. ZFNs can be used in any plant species amenable to DNA delivery; our results therefore establish a newstrategy for plant genetic manipulation in basic science and agricultural applications.