Precise Genome Modification in the Crop Species Zea Mays Using Zinc-Finger Nucleases.

Agricultural biotechnology is limited by the inefficiency and unpredictability of conventional random mutagenesis and transgenesis.  Here we report a broadly applicable, versatile alternative: the use of designed zinc-finger nucleases (ZFNs) that induce double-stranded breaks (DSB) at their target locus.  We have used ZFNs to modify endogenous loci in plants of the crop species Zea mays, and show that ZFNs designed to cleave a maize gene disrupt their target via DSB repair by error-prone non-homologous end-joining.  We further demonstrate that simultaneous expression of ZFNs and delivery of a simple heterologous donor molecule leads to high frequency targeted addition of an herbicide-tolerance gene at the intended locus.  Modified maize plants transmit these genetic changes to the next generation.  Insertional disruption of one target locus, ZmIPK1, results in both herbicide tolerance and alterations of the inositol phosphate profile in developing seeds.  ZFNs can be utilized in any plant species amenable to DNA delivery; therefore our results establish a new paradigm in plant genetic manipulation for basic science and agricultural applications.