Phylogeny analysis of Indian strains of Rhizoctonia solani isolated from chickpea and development of sequence characterized amplified region (SCAR) marker for detection of the pathogen

Abstract

Wet root rot caused by Rhizoctonia solani is one of the important diseases in chickpea worldwide. In the present study, 10 random amplified polymorphic DNA (RAPD) primers were used to assess the molecular diversity of 50 chickpea isolates of R. solani. There was a great diversity among the isolates studied and was in the range of 52 to 93%. The isolates were highly variable in aggressiveness and caused up to 100% wet root rot incidence in chickpea. Accurate detection and identification of plant pathogens are fundamental to plant pathogen diagnostics and management. Therefore, a polymerase chain reaction (PCR) assay was developed for accurate and sensitive detection of R. solani from mycelial DNA and infected chickpea plants. RAPD primer OPA 11 consistently amplified ≈1700 base pairs (bp) product in PCR only from the DNA of R. solani isolated from chickpea. The common DNA fragment was sequenced and used to design a pair of oligonucleotide primers amplifying 285 bp sequence characterized amplified region (SCAR). The specificity of the SCAR primers was evaluated. The detection sensitivity of R. solani was 0.5 ng for the genomic DNA and 5 ng for the DNA extracted from infected chickpea root samples. Also, SCAR primer was validated with Q-PCR to detect and quantify R. solani upto 1 pg from infected chickpea root samples. These new SCAR marker are useful for early detection and quantification of wet root rot pathogen in chickpea.