Genetic mapping of disease resistance gene analogs from the Italian ryegrass (Lolium multiflorum Lam.) genome

Abstract

Genomic DNA sequences homologous to the nucleotide binding site (NBS)-leucine-rich repeat (LRR) genes were mapped for Italian ryegrass (Lolium multiflorum Lam.) by using previously published disease resistance gene analogs (RGAs). The RGAs were cloned by means of a nested polymerase chain reaction (PCR) approach with degenerate primers designed from conserved regions of the NBS domain of plant disease resistance genes (R-genes), and primer pairs for specific sequence-tagged sites (STS) were subsequently designed for efficient generation of the RGA fragments. To map the RGA clones on the Italian ryegrass genetic map, an F1 mapping family was used for the detection of restriction-site polymorphisms of the RGA fragments. Out of 50 RGA fragments, 11 detected genetic polymorphism in the F1 family. Polymorphic marker loci data were used for linkage analysis with JoinMap version 3.0. The linkage analysis revealed that 10 of the 11 RGAs were mapped in seven linkage groups constructed with amplified fragment length polymorphism (AFLP), simple sequence repeat (SSR), and expressed sequence tag (EST)-derived cleaved amplified polymorphic sequence (CAPS) markers. This information will be useful for the development of new genetic markers linked to genes associated with disease resistance.