밤나무 잉크병균, Phytophthora katsurae의 검출을 위한 Duplex PCR

초록

Phytophthora katsurae is a fungal pathogen responsible for chestnut ink disease. We designed two duplex primer sets (SOPC 1F/1R+KatI 3F/5R, SOPC 1-1F/1-1R+KatI 3F/5R) to detect P. katsurae. SOPC 1F/1R and SOPC 1-1F/1-1R primer pairs were designed for sequence characteristic amplification regions (SCAR)marker, and KatI 3F/5R primer pair was used for P. katsurae-specific primer designed from internal transcribed spacer (ITS) region. To assess the sensitivity of duplex PCR, genomic DNA was serially diluted 10-fold to make the final concentrations from 1 mg/ml to 1 ng/ml. The sensitivity for two primer sets were 1μg/ml and 100 ng/ml, respectively. To find detection limits for zoospores of P. katsurae, each zoospore suspension was serially diluted 10-fold to make the final concentrations from 1× 106 to 1 × 102 cells/ml, and then DNA was extracted. The limits of detection for all of two primer sets were 1 × 105 cells/ml. All of two primer sets were specific to P. katsurae in PCR detection and did not produce any P. katsurae-specific PCR amplicons from other 16 Phytophthora species used as the control. This study shows that duplex PCR using two primer sets might be a useful tool for rapid and efficient detection of P. katsurae.

키워드