Development of <i>Aegilops comosa</i> and <i>Aegilops caudata</i>‐specific molecular markers and fluorescence <i>in situ</i> hybridization probes based on specific‐locus amplified fragment sequencing

As tertiary gene pools of wheat, Aegilops comosa and Ae. caudata contain many excellent genes/traits and gradually become important and noteworthy wild resources for wheat improvement worldwide. However, the lack of molecular markers and cytological probes with good specificity and high sensitivity limits the development and utilization of Triticum aestivum-Ae. comosa (Ta. Aeco)/Ae. caudata (Ta. Aeca) introgression lines. Using specific-locus amplified fragment sequencing, two Ae. comosa and one Ae. caudata accessions, Chinese Spring, and three Ta. Aeco and Ta. Aeca introgression lines each were sequenced to develop new molecular markers and cytological probes. After strict sequence comparison and verification in different materials, a total of 39 molecular markers specific to three chromosomes in Ae. comosa (nine, seven, and 10 markers for 1M, 2M, and 7M, respectively) and Ae. caudata (two, six, and five markers for 3C, 4C, and 5C, respectively) and 21 fluorescence in situ hybridization (FISH) probes (one centromeric probe with signals specific to the M chromosomes, two centromeric probes with signals in all the tested genomes, and six, eight, and four FISH probes specific to the M, C, and M, C, and U chromosomes, respectively) were successfully exploited. The newly developed molecular markers and cytological probes could be used in karyotype studies, centromere evolutionary analyses of Aegilops, and had the ability to detect the fusion centromeres and small-fragment translocations in introgression lines.