Induction of Stable Epigenetic Gene Silencing in Plants Using a Virus Vector

Protocol
First Online:
Part of the Methods in Molecular Biology book series (MIMB, volume 1287)

Abstract

Gene silencing through transcriptional repression can be induced by double-stranded RNA targeted to a gene promoter, a process known as RNA-mediated transcriptional gene silencing (TGS). This phenomenon is associated with epigenetic changes involving cytosine methylation of the promoter. Plant virus vectors have been used to induce RNA-mediated TGS. Here, we describe methods relevant to the induction of epigenetic changes and RNA-mediated TGS in plants using a virus vector, which include inoculation of recombinant virus, detection of short interfering RNAs, bisulfite sequencing analysis, and nuclear run-on transcription assay.

Key words

Epigenetic changes RNA-directed DNA methylation (RdDM) RNA silencing Transcriptional gene silencing (TGS) Virus-induced gene silencing (VIGS) Virus vector 
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Notes

Acknowledgments

We thank the coauthors of our papers cited in this protocol for their cooperation. This work was supported by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan.

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Research Faculty of AgricultureHokkaido UniversitySapporoJapan

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