Leymus flavescens (Scribner&Smith) Pilger, yellow wild rye, is a long-lived, strongly rhizomatous, tetraploid (2n54x528) perennial grass of the tribe Triticeae distributed throughout centralWashington, eastern Oregon, and the Snake River plains of Idaho. Our objectives were (1) to describe chromosome pairing and fertility in F1 hybrids between L. flavescens and North American tetraploids (2n54x528) L. triticoides and L. cinereus and Eurasian tetraploids L. secalinus, L. racemosus, and L. alaicus subsp. karataviensis and (2) to utilize genome-specific random amplified polymorphic DNA (RAPD) markers to verify the genomic composition of L. flavescens. The hybrids L. flavescens # L. triticoides (NsNsXmXm), L. flavescens # L. secalinus (Ns- NsXmXm), L. flavescens # L. racemosus (NsNsXmXm), L. flavescens # L. cinereus (NsNsXmXm), and L. flavescens# L. alaicus subsp. karataviensis (NsNsXmXm) averaged 13.9, 13.8, 13.6, 13.1, and 11.9 bivalents per cell, respectively. Genome-specific RAPD assay indicates that L. flavescens has the Ns genome but lacks the St genome from the genus Pseudoroegneria and the H genome from the genus Hordeum. On the basis of the bivalent chromosome pairing frequency in the F1 hybrids of L. flavescens, the genomic formula of L. flavescens is NsNsXmXm. The presence of the Ns genome was verified by molecular characterization.