Journal
PLANT PATHOLOGY
Volume 68, Issue 5, Pages 878-888Publisher
WILEY
DOI: 10.1111/ppa.13015
Keywords
EU1 and EU2 lineages; NA1; NA2; sudden oak death; TaqMan probes; unique genes
Categories
Funding
- Genome Canada's Large-Scale Applied Research Program (LSARP) [2112, 10106]
- Genome Canada's Genomic Application Partnership Program [GAPP 6102]
- Genome B.C.
- Genome Quebec
- Canadian Food Inspection Agency
- FPInnovations
- Natural Resources Canada
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Early detection provides the best way to prevent introduction and establishment of alien plant pathogens. Amplification of DNA by PCR has revolutionized the detection and monitoring of plant pathogens. Most of those assays rely on the amplification of a fraction of the genome of the targeted species. With the availability of whole genomes for a growing number of fungi and oomycetes it is becoming possible to compare genomes and discover regions that are unique to a target organism. This study has applied this pipeline to develop a set of hierarchical TaqMan real-time PCR detection assays targeting DNA of all four Phytophthora ramorum lineages, and a closely related species, P. lateralis. Nine assays were generated: three targeting DNA of all P. ramorum lineages, one for each lineage of P. ramorum, one for P. lateralis and one targeting DNA of P. ramorum and P. lateralis. These assays were very accurate and sensitive, ranging from 98.7% to 100% detection accuracy of 2-10 gene copies of the targeted taxa from pure cultures or inoculated tissues. This level of sensitivity is within the lowest theoretical limit of detection of DNA. It is expected that these assays will be useful because of their high level of specificity and the ease with which they can be multiplexed because of the inherent flexibility in primer and probe design afforded by their lack of conservation in non-target species.
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