Study on DNA polymorphism in populations of Rhizoctonia solani anastomosis group 4 using rDNA RFLP

Ribosomal DNA (rDNA) sequences have been widely used to study the phylogenetic relationships in different fungi. Fungal nuclear rRNA genes are arranged as tandem repeats with several hundred copies per genome. These spacer regions are considerably more variable than the subunit sequences and have been widely used in studies on the relationships among species within a single genus or among intraspecific populations. To evaluate the polymorphism between 18S and 28S genes, 28 isolates of Rhizoctonia solani anastomosis group 4 were examined. Genomic DNA was extracted from the isolates and prepared for PCR reaction. The amplification was performed using internal transcribed spacer (ITS) ITS1 and ITS4 primers. A DNA fragment of 700 bp in size was detected in PCR products of all tested isolates. To assess existence of any further polymorphism in the ITS region, the PCR products were digested with restriction endonucleases. Although there were restriction sites for XbaI, HaeIII, BamHI, HindIII, SacI, PstI, and TaqI endonucleases, there were no restriction sites for NdeI, XhoI, HincII, HinfI, EcoRI and XhoI endonucleases. The endonuclease HincII recognized a restriction site on PCR products that discriminated the isolates belonging to AG4-HGII form isolates of AG4-HGI. Based on the results, it has been concluded that AG-4 isolates of Rhizoctonia solani wereheterogenic.