The species of genus Aegilops were not domesticated and exhibit large genetic and phenotypic diversity. The U- and Mgenome species Ae. umbellulata (2n=2x=14; UU) and Ae. comosa (2n=2x=14, MM) and their allotetraploid hybrids are rich sources of useful genes and alleles for wheat improvement. The effective gene transfer from these Aegilops species via interspecific hybridization requires the identification of U and M chromosomes in the wheat genetic background. In situ hybridization with fluorescently labelled satellite DNA probes is a powerful method for individual chromosome identification and monitoring alien gene introgression into wheat. In the present work, we identified satellite repeats using sequence data of flow sorted Aegilops chromosomes and investigated the ability to use them as probes for FISH for the characterization of U and M chromosomes. The chromosomes 1U, 3U, 4U, 5U, 6U, and 7U and chromosome arms 2US and 2UL of Ae. umbellulata were flow sorted and sequenced by Illumina. Out of the "26 Gbp sequence data obtained per chromosome, "316.6 Mbp/chromosome were assembled to cover "40% of the estimated U genome size. Graph-based clustering of 1,420,161 reads resulted in 34,745 clusters where the most abundant repeats were different types of LTR retrotransposons: Ty3/gypsy elements (34.52%), Ty1/copia elements (9.08%) and putative non-annotated LTR retroelements (5.00%). Putative tandem repeats were identified based on graph layout and Dotter analysis. Tandem repeats were PCR-amplified using genomic DNA of Ae. umbellulata as template with specific oligo-primers and labelled with biotin. Five out of the 12 satellite repeats showed characteristic signals on the U and M genome chromosomes of diploid Ae. umbellulata and Ae. comosa. The repeats showed specific signals on one chromosome (CL199, CL70), on three chromosomes (CL174) or on six chromosomes (CL191) of the diploid species. The various combinations of these satellite repeats with the probes (pSc119.2, Afa family, pTa71) previously used for the karyotypic analysis of Triticum/Aegilops will facilitate the cytogenetic selection of wheat–Aegilops addition, substitution or translocation lines.