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Host-Induced Gene Silencing (HIGS) for Elucidating Puccinia Gene Function in Wheat

Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 1848)

Abstract

Biotrophic fungi (Puccinia spp.) cause devastating diseases of wheat and other cereal species globally. The function of large repertories of genes from Puccinia spp. still needs to be discovered to understand the infection process of these obligate parasites, eventually to protect plants from rust diseases. Functional analysis of targeted genes is challenging due to the inherent difficulties with culturing the fungus and transforming the host. RNA interference (RNAi) is a conserved gene regulation process in eukaryotes and known to be a powerful genetic tool in plant biotechnology. More recently, host-induced gene silencing (HIGS) has been developed to assess pathogen gene function in plants. HIGS is an RNAi-based process where double stranded RNA (dsRNA) homologous to a pathogen gene can be expressed in a plant to induce targeted silencing of the pathogen gene. Here we described a detailed HIGS protocol for functional analysis of rust genes from Puccinia species in cereals. As an example we describe an experiment silencing the tryptophan 2-monooxygenase gene (Pgt-IaaM) from Puccinia graminis f. sp. tritici (Pgt) that is involved in virulence to wheat.

Key words

HIGS Puccinia Pathogenicity Barley mosaic stripe virus Pgt-IaaM 
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Notes

Acknowledgments

This work was funded by National Institute of Food and Agriculture to S.H. (Grant No. 2010-65108-20568).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Plant PathologyWashington State UniversityPullmanUSA

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