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United States Department of Agriculture

Agricultural Research Service

Title: Real-Time Fluorescent Pcr Detection of the Sudden Oak Death Pathogen Phytophthora Ramorum

Authors
item Tooley, Paul
item Martin, Frank
item Carras, Marie
item Frederick, Reid

Submitted to: Mycological Society of America
Publication Type: Abstract Only
Publication Acceptance Date: April 1, 2004
Publication Date: August 1, 2004
Citation: Mycologia 55:37

Technical Abstract: Phytophthora ramorum causes sudden oak death, a disease which has killed many oaks in California and which also threatens the nursery industry due to its ability to cause foliar infection on rhododendrons and many other ornamentals. A real-time PCR detection method for P. ramorum was developed based on mitochondrial DNA sequence and using an ABI Prism 7700 Sequence Detection System (Taqman). Primers and probes were also developed for detecting P. pseudosyringae, a newly described species which causes symptoms similar to those of P. ramorum on certain hosts. The Phytophthora-specific primer-probe system was used in a multiplex assay with a plant primer-probe system utilizing the dyes FAM and CAL Orange to allow plant DNA to serve as a positive control in each reaction. With the FAM dye system, detection of genomic P. ramorum DNA was achieved down to 1 fg of DNA. P. ramorum could be detected from inoculated Rhododendron cv. 'Cunningham's White' (12 mg fresh weight tissue) down to a 10 EE -5 dilution. In a blind test, the method also successfully detected P. ramorum in California field samples previously found to be positive based on conventional PCR and isolation of the pathogen in pure culture. This system should prove useful in allowing rapid, sensitive and specific detection of the sudden oak death pathogen on ornamentals and other host species.

Last Modified: 11/22/2014
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