关键词: 大豆疫霉菌, SSR标记, 分子检测, 巢式PCR

Abstract:

【Objective】 This study was aimed at developing a rapid and accurate molecular technique for detection of Phytophthora sojae and diagnosis of soybean Phytophthora root rot at early stage. 【Method】 Based on the SSR marker Psc239 of P. sojae, two primer pairs, Psc239EF/Psc239ER and Psc239F/Psc239R, were designed specifically to amplify DNA from P. sojae by nested PCR. 【Result】 The primer pairs and nested PCR assay were assessed for specificity using DNA from 36 isolates of P. sojae and 25 isolates of other oomycetes and fungi, Psc239EF/Psc239ER, Psc239F/Psc239R and nested PCR could amplify 519 bp, 242 bp and 242 bp fragments only from P. sojae, respectively, no amplification product was observed from other oomycetes and fungi. In conventional PCR, the sensitivity of detection was of 50 pg?μl-1 P. sojae DNA for both primer pairs, in the nested PCR the detection limit was 50 f g?μl-1. The detection limit for oospores of P. sojae in soil was of 0.4 oospores in 1g soils by the nested PCR. The nested PCR assay was also used to detect DNA extracted from infected soybean tissues and soil samples. 【Conclusion】 The nested PCR assay based on SSR marker Psc239 could be used to rapidly detect P. sojae in soil and diagnose Phytophthora root rot on soybean.

Key words: Phytophthora sojae, SSR marker, molecular detection, nested PCR