Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2005 Mar 1;6(2):153-63.
doi: 10.1111/j.1364-3703.2005.00272.x.

A method for double-stranded RNA-mediated transient gene silencing in Phytophthora infestans

Stephen C Whisson  1 , Anna O Avrova, Pieter VAN West, John T Jones
Affiliations

A method for double-stranded RNA-mediated transient gene silencing in Phytophthora infestans

Stephen C Whisson et al. Mol Plant Pathol. .

Abstract

SUMMARY Gene silencing, triggered by double-stranded RNA (dsRNA), has proved to be a valuable tool for determining and confirming the function of genes in many organisms. For Phytophthora infestans, the cause of late blight on potato and tomato, gene silencing strategies have relied on stable transformation followed by spontaneous silencing of both the endogenous gene and the transgene. Here we describe the first application of transient gene silencing in P. infestans, by delivering in vitro synthesized dsRNA into protoplasts to trigger silencing. A marker gene, gfp, and two P. infestans genes, inf1 and cdc14, both of which have been silenced previously, were selected to test this strategy. Green fluorescent protein (GFP) fluorescence was reduced in regenerating protoplasts up to 4 days after exposure to gfp dsRNA. A secondary reduction in expression of all genes tested was not fully activated until 12-17 days after introduction of the respective homologous dsRNAs. At this time after exposure to dsRNA, reduced GFP fluorescence in gfp dsRNA-treated lines, and reduced INF1 production in inf1 dsRNA-treated lines, was observed. Introduction of dsRNA for the stage-specific gene, cdc14, yielded the expected phenotype of reduced numbers of sporangia when cdc14 expression was significantly reduced compared with control lines. Silencing was shown to be sequence-specific, because analysis of inf1 expression in gfp-silenced lines yielded wild-type levels of gene expression. This report shows that transient gene silencing can be used to generate detectable phenotypes in P. infestans and should provide a high-throughput tool for P. infestans functional genomics.

PubMed Disclaimer

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

LinkOut - more resources