Regions of amino acid conservation in the NBS domain of NBS-LRR resistance proteins facilitated the PCR isolation of eight resistance gene analog (RGA) sequences from genomic DNA of rice, barley, and Aegilops tauschii. These clones and other RGAs previously isolated from maize, rice, and wheat were assigned to 13 classes by DNA-sequence comparison and by their patterns of hybridisation to restricted barley DNA. Using a doubled-haploid mapping population, probes from 12 RGA classes were used to map 17 loci in the barley genome. Many of these probes have been used for mapping in wheat, and the collective data indicate that the positions of orthologous RGAs are conserved between barley and wheat. RGA loci were identified in the vicinity of barley leaf rust resistance loci Rph4, Rph7, and Rph10. Recombinants were identified between RGA loci and Rph7 and Rph10, while a cluster of RGA sequences detected by probe 5.2 cosegregated with Rph4 in 55 F2 lines.