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

Nature volume 459, pages 437–441 (2009)Cite this article

Abstract

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 of designed 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 of ZFNs and delivery of a simple heterologous donor molecule leads to precise targeted addition of an herbicide-tolerance gene at the intended locus in a significant number of isolated events. ZFN-modified 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 new strategy for plant genetic manipulation in basic science and agricultural applications.

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Figure 1: ZFNs designed to target Z. mays IPK1 induce DSBs at the endogenous locus.
Figure 2: Targeted gene addition to IPK1 in maize.
Figure 3: ZFN-mediated gene disruption of Z. mays IPK1 is stable and heritable.

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Acknowledgements

The authors wish to thank numerous colleagues at Dow AgroSciences and Sangamo BioSciences for discussions regarding this project, the DAS Gamma group for statistics support, Sangamo’s production group for technical support and A. Klug for a critical reading of the manuscript. We are also grateful to W. Kleschick, D. Kittle and E. Lanphier for encouragement.

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  1. Dow AgroSciences, 9330 Zionsville Road, Indianapolis, Indiana 46268, USA ,

    Vipula K. Shukla, Sarah E. Worden, Jon C. Mitchell, Nicole L. Arnold, Gao Zhifang, David McCaskill, Matthew A. Simpson, Beth Blakeslee, Scott A. Greenwalt & Holly J. Butler

  2. Sangamo BioSciences, Point Richmond Tech Center, 501 Canal Boulevard, Suite A100, Richmond, California 94804, USA ,

    Yannick Doyon, Jeffrey C. Miller, Russell C. DeKelver, Erica A. Moehle, Sunita Gopalan, Xiangdong Meng, Vivian M. Choi, Jeremy M. Rock, Ying-Ying Wu, George E. Katibah, Sarah J. Hinkley, Lei Zhang, Edward J. Rebar, Philip D. Gregory & Fyodor D. Urnov

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Correspondence to Vipula K. Shukla.

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The authors are current or recent employees of Dow AgroSciences, LLC, a wholly owned subsidiary of the Dow Chemical Company, or Sangamo BioSciences. These companies may gain or lose financially through this publication. The authors may also own stocks or shares in companies including the Dow Chemical Company and Sangamo BioSciences, that may gain or lose financially through publication of this article. Several authors are inventors on patents or patent applications whose value may be affected by this publication.

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Shukla, V., Doyon, Y., Miller, J. et al. Precise genome modification in the crop species Zea mays using zinc-finger nucleases. Nature 459, 437–441 (2009). https://doi.org/10.1038/nature07992

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