Two clones were isolated from oat (A vena sativa cv. Hinoat) genomic DNA. Restriction analysis and cross-hybridization studies revealed that these clones represent two different families of repeated DNA. Single accessions of 10 Avena species, representing three ploidy levels (2x, 4x, and 6x; x = 7) and comprising all available genomic constitutions (AA, CC, AABB, AACC, and AACCDD), were analyzed with these probes. Differences were observed among single representatives of the 10 species in the relative abundance of each probe and between the probes in relative abundance in each species. In slot-blot experiments, for each of the two probes, hybridization intensities were independent of genome composition. Southern blots of HindIII DNA digests from the 10 species, using each of the two probes, allowed classification of these species into two groups: the three diploid species with a CC genomic constitution had one specific pattern and the remaining seven species having different genomic constitutions (AA, AABB, AACC, AACCDD, the A genome being in common) had a different pattern. Therefore, these probes will distinguish the C-genome diploid species from the diploid and polyploid species carrying the A genome.