Development and application of duplex and triplex assays for simultaneous detection of resistance genes to leaf rust, Fusarium head blight, powdery mildew, Septoria tritici blotch, eyspot, stem rust and yellow rust in wheat

Resistance breeding is a widely promoted strategy for minimizing yield losses in wheat caused by various fungal diseases, including leaf rust (Puccinia triticina; Lr genes), Fusarium head blight (Fusarium spp.; Fhb), powdery mildew (Blumeria graminis; Pm), Septoria tritici blotch (Septoria tritici; Stb), eyespot (Oculimacula spp.; Pch-previously known as Pseudocercosporella herpotrichoides), stem rust (Puccinia graminis f. sp. tritici; Sr) and yellow rust (Puccinia striiformis; Yr). Understanding the prevalence of resistance genes in currently cultivated European varieties is crucial for their effective utilization in breeding programs. In this study, we developed 11 duplex and 13 triplex PCR assays for the simultaneous identification of diverse allelic combinations of disease resistance genes. Selected assays were used to analyze 70 European wheat varieties for the presence of 15 molecular markers associated with 12 resistance genes. The analyses identified diverse resistance gene combinations. The optimized multiplex PCR methods significantly reduce cost and time of analysis, making them valuable tools for marker-assisted selection (MAS) in wheat breeding programs.