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Improving a Quantitative Trait in Rice by Multigene Editing with CRISPR-Cas9

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Rice Genome Engineering and Gene Editing

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2238))

Abstract

CRISPR-Cas9 system is one sequence-specific nuclease (SSN) that has several advantages over zinc finger nuclease (ZFN) and transcription activator-like effector nuclease (TALEN), such as multiplex genome editing. With multiplex genome editing, CRISPR-Cas9 becomes a preferred SSN to edit many quantitative trait loci (QTL) simultaneously for trait improvement in major crops. A multiplexed CRISPR system is also important for deletion of a large fragment within a chromosome, analysis of the function of gene families, exon exchange, gene activation, and repression. Therefore, assembly of several single guide RNAs (sgRNAs) into one binary vector is the main step in multigene editing by CRISPR-Cas9. Different vector construction methods have been practiced including Golden Gate assembly. This chapter provides a detailed protocol for the construction of a T-DNA binary vector for expressing Cas9 and three sgRNAs for simultaneous targeting of three QTL genes for improving seed trait in rice.

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Acknowledgments

This work was supported by grants including the National Science Foundation of China (31771486), the Sichuan Youth Science and Technology Foundation (2017JQ0005), the National Transgenic Major Project (2018ZX08022001-003) and the Science Strength Promotion Program of UESTC to Y.Z., and the National Science Foundation Plant Genome Research Program (IOS-1758745) and USDA-NIFA Biotechnology Risk Assessment Research Program (2018-33522-28789) to Y.Q.

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Authors and Affiliations

  1. Department of Biotechnology, School of Life Science and Technology, Center for Informational Biology, University of Electronic Science and Technology of China, Chengdu, China

    Yesuf Teslim Yimam, Jianping Zhou, Sayed Abdul Akher, Xuelian Zheng & Yong Zhang

  2. Department of Plant Science and Landscape Architecture, University of Maryland, College Park, MD, USA

    Yiping Qi

  3. Institute for Bioscience and Biotechnology Research, University of Maryland, Rockville, MD, USA

    Yiping Qi

Authors
  1. Yesuf Teslim Yimam
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  2. Jianping Zhou
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  3. Sayed Abdul Akher
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  4. Xuelian Zheng
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  5. Yiping Qi
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  6. Yong Zhang
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Corresponding author

Correspondence to Yong Zhang .

Editor information

Editors and Affiliations

  1. Synthetic Biology, Biofuel and Genome Editing R&D, Reliance Industries Ltd, Navi Mumbai, Maharashtra, India

    Anindya Bandyopadhyay

  2. Crop Improvement and Genetics Research Unit, USDA-ARS, Albany, CA, USA

    Roger Thilmony

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Yimam, Y.T., Zhou, J., Akher, S.A., Zheng, X., Qi, Y., Zhang, Y. (2021). Improving a Quantitative Trait in Rice by Multigene Editing with CRISPR-Cas9. In: Bandyopadhyay, A., Thilmony, R. (eds) Rice Genome Engineering and Gene Editing. Methods in Molecular Biology, vol 2238. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1068-8_13

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  • DOI: https://doi.org/10.1007/978-1-0716-1068-8_13

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1067-1

  • Online ISBN: 978-1-0716-1068-8

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