Development of PCR based diagnostic techniques for the two mating types ofPyrenopeziza brassicae (light leaf spot) on winter oilseed rape (Brassica napus ssp oleifera)

Two sets of PCR primers were designed from DNA sequence which flanked the 3 ' end of the Pyrenopeziza brassicae mating type idiomorphs (MAT-1 and MAT-2 ). The first set of primers facilitated the amplification of a 750 bp produ ct from genomic DNA of 50 isolates of P. brassicae, which included both mat ing types. The 750 bp PCR product was also amplified from DNA extracted fro m Brassica napus tissue infected with P. brassicae. The 750 bp PCR product was not produced when these diagnostic primers were used in PCR reactions c ontaining DNA from a number of other fungal species, including fungal patho gens of oilseed rape and Tapesia yallundae, which is closely related to P. brassicae. The second set of primers were designed to amplify different siz ed products from the two mating types of P. brassicae. These mating type sp ecific primers facilitated the amplification of a 440 bp product from all M AT-1 isolates tested and a 535 bp product from 30% of the MAT-2 isolates te sted. The value of molecular diagnostic techniques for the identification o f P. brassicae in oilseed rape crops, for distinguishing between mating typ es of the fungus and in disease forecasting is discussed. (C) 1999 Academic Press.