利用荧光定量PCR检测禾谷丝核菌的相对生物量

Relative quantification of biomass of Rhizoctonia cerealis using real-time PCR

小麦纹枯病是以禾谷丝核菌(Rhizoctonia cerealis)侵染为主的小麦土传病害。为建立检测禾谷丝核菌在寄主小麦(Triticum aestivum)中的相对生物量的可靠方法,促进小麦抗纹枯病机制的研究,本研究克隆了禾谷丝核菌肌动蛋白基因RcActin的部分(3′端)cDNA,并设计了RcActin的特异引物。该引物不仅能区分禾谷丝核菌与寄主小麦,还能区分全蚀病菌(Gaeumannomyces graminis)、根腐病菌(Bipolaris sorokiniana)和立枯丝核菌(Rhizoctonia solani)等常见小麦土传病害的病原菌,表明该引物能用于小麦纹枯病的分子检测,也能用于相对表达量的测定。利用相对定量法,以RcActin相对于寄主管家基因的相对表达量作为禾谷丝核菌相对生物量的指标,结果表明,此方法能准确反映禾谷丝核菌在寄主中的相对生物量和对小麦纹枯病抗性程度进行快速鉴定。

Wheat sharp eyespot, caused by Rhizoctonia cerealis, is a devastating soil-borne disease. In order to quantify the relative biomass of R. cerealis and propel the study about mechanism of resistance of wheat against R. cerealis, we had cloned the partial cDNA of actin gene from R. cerealis （named as RcActin）. Then the specific primers of RcActin were designed for diagnosis of sharp eyespot and expression analysis of RcActin. The primers could specifically distinguish R. cerealis from wheat, Gaeurnannomyces graminis, Bipolaris sorokiniana and Rhizoctonia solani. We had proofed that the RcActin expression level can be used as an indicator of the fungal relative biomass and evaluation of resistant degree against R. cerealis in wheat.