B Section B : B-7 ; Development of Rapid and Accurate PCR Assay for Phytophthora ramorum in Blueberry Plant

Phytophthora ramorum is the causal agent for the disease commonly called sudden oak death. In addition, the pathogen also causes destructive symptoms on a wide-and ever-increasing-range of woody and herbaceous hosts. Thus, spread of this pathogen is still a potential threat in Korea. For these reasons, the detection of P. ramorum is important to prevent pathogen spread, the quarantine authorities of in Korea has enforced strict measures, regulating import of host plant materials. To develop a rapid and accurate PCR assay for P. ramorum from suspicious plant, newly developed species-specific primers based on the DNA sequences of Ypt1 gene (ras-related protein) amplified 140 to 190bp PCR products for P. ramorum. These primer sets showed no reaction in test against other Phytophthora species as well as phytopathogenic fungi, respectively. The detection limit of species-specific primer sets was 1 to 10pg/μl in conven-tional PCR. The nested PCR showed a detection limit about 10 times better than a single round PCR. The PCR assay using specific primer sets and a NaOH DNA extraction method was able to detect quickly P. ramorum from artificially inoculated blueberry roots on a Phytophthora semi-selective medium (COA involves antibiotics; Ampicillin, Rifampicin, Nystatin) in effendorf tube. This PCR assay can be utilized for rapid/accurate detection and identification of P. ramorum practically for the plant quarantine.