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A Sensitized Genetic Screen to Identify Novel Components and Regulators of the Host Antiviral RNA Interference Pathway

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Book cover Antiviral Resistance in Plants

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2028))

Abstract

RNA interference (RNAi) acts as a natural defense mechanism against virus infection in plants and animals. Much is known about the antiviral function of the core RNAi pathway components identified mostly by genetic screens based on specific RNAi of cellular mRNAs. Here we describe a sensitized genetic screening system for the identification of novel components and regulators in the antiviral RNAi pathway established in the model plant species Arabidopsis thaliana. Our genetic screen identifies antiviral RNAi (avi)-defective Arabidopsis mutants that develop visible disease symptoms after infection with CMV-∆2b, a Cucumber mosaic virus mutant deficient in the expression of its viral suppressor of RNAi. Loss of RNAi suppression renders CMV-∆2b highly susceptible to antiviral RNAi so that it replicates to high levels and induces disease development only in avi mutants. This chapter provides the methods for the propagation of CMV-∆2b, preparation of the mutant plants for virus inoculation, identification and characterization of avi mutants, and cloning of the genes responsible for the mutant phenotype by either the genetic linkage to T-DNA insertion or a mapping-by-sequencing approach.

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Acknowledgments

This project was supported by grants from the US Department of Agriculture Research Service (6659-22000-025), the US-Israel Binational Agricultural Research and Development Fund, National Institutes of Health (R01AI52447 and GM94396), by the Agricultural Experimental Station of the University of California, Riverside (to S.-W.D.)

Author information

Authors and Affiliations

  1. Vector-Borne Virus Research Center, Haixia Institute of Science and Technology, College of Plant Protection, Fujian Agriculture and Forestry University, Fuzhou, Fujian, People’s Republic of China

    Zhongxin Guo

  2. State Key Laboratory of Agro-Biotechnology, College of Biological Sciences, China Agricultural University, Beijing, People’s Republic of China

    Xian-Bing Wang

  3. Department of Microbiology and Plant Pathology and Center for Plant Cell Biology, University of California, Riverside, CA, USA

    Wan-Xiang Li & Shou-Wei Ding

Authors
  1. Zhongxin Guo
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  2. Xian-Bing Wang
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  3. Wan-Xiang Li
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  4. Shou-Wei Ding
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Corresponding authors

Correspondence to Xian-Bing Wang or Shou-Wei Ding .

Editor information

Editors and Affiliations

  1. Faculty of Agriculture, Ehime University, Matsuyama, Ehime, Japan

    Kappei Kobayashi

  2. Faculty of Agriculture, Ehime University, Matsuyama, Ehime, Japan

    Masamichi Nishiguchi

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Guo, Z., Wang, XB., Li, WX., Ding, SW. (2019). A Sensitized Genetic Screen to Identify Novel Components and Regulators of the Host Antiviral RNA Interference Pathway. In: Kobayashi, K., Nishiguchi, M. (eds) Antiviral Resistance in Plants. Methods in Molecular Biology, vol 2028. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9635-3_12

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  • DOI: https://doi.org/10.1007/978-1-4939-9635-3_12

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-9634-6

  • Online ISBN: 978-1-4939-9635-3

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