Abstract:
Sequence characterized amplified region (SCAR) markers are one of the most effective and accurate tools for microbial
identification. In this study, we applied SCAR markers for the rapid and accurate detection of Phytophthora katsurae, the casual
agent of chestnut ink disease in Korea. In this study, we developed seven SCAR markers specific to P. katsurae using random
amplified polymorphic DNA (RAPD), and assessed the potential of the SCAR markers to serve as tools for identifying P. katsurae.
Seven primer pairs (SOPC 1F/SOPC 1R, SOPC 1-1F/SOPC 1-1R, SOPC 3F/SOPC 3R, SOPC 4F/SOPC 4R, SOPC 4F/SOPC 4-1R,
SOPD 9F/SOPD 9R, and SOPD 10F/SOPD 10R) from a sequence derived from RAPD fragments were designed for the analysis
of the SCAR markers. To evaluate the specificity and sensitivity of the SCAR markers, the genomic DNA of P. katsurae was serially
diluted 10-fold to final concentrations from 1 mg/mL to 1 pg/mL. The limit of detection using the SCAR markers ranged from
100 μg/mL to 100 ng/mL. To identify the limit for detecting P. katsurae zoospores, each suspension of zoospores was serially
diluted 10-fold to final concentrations from 10 × 105 to 10 × 101 zoospores/mL, and then extracted. The limit of detection by
SCAR markers was approximately 10 × 101 zoospores/mL. PCR detection with SCAR markers was specific for P. katsurae, and did
not produce any P. katsurae-specific PCR amplicons from 16 other Phytophthora species used as controls. This study shows that
SCAR markers are a useful tool for the rapid and effective detection of P. katsurae.
