RNA-seq facilitates development of chromosome-specific markers and transfer of rye chromatin to wheat

Transcriptome shotgun sequencing (RNA-seq) provides an abundant resource for developing molecular markers specifically related to functional genes. In this study, transcriptomes of Chinese rye cultivar Jingzhouheimai (JZHM) challenged with powdery mildew pathogen Bgt (Blumeria graminis f. sp. tritici) were obtained and used to develop expressed sequence tag (EST)-based simple sequence repeat (SSR) and sequence-tagged site (STS) markers. A total of 866 primer sets for EST-SSRs and STSs were designed, from which we developed 401 rye-specific markers. The highest level of polymorphism was observed in EST-SSRs (56.73%) followed by STS2 (49.07%) designed via rye-specific contigs and STS1 (35.90%) primers designed from upregulated contigs. Genotyping with newly developed markers along with cytogenetic techniques allowed us to identify nine wheat alien chromosome lines from the cross of Zhoumai 18/Jinghui 1 (Jinghui 1 is an amphiploid of wheat landrace Huixianhong and JZHM) carrying rye chromosome 6R or 6R segments of different length, which permitted preliminary location of the powdery mildew resistance gene PmJZHM6RL and 12 specific markers to 6RL FL 0.51–1.0. 5R-specific markers and genomic in situ hybridization/fluorescence in situ hybridization detected MtA5RL and T5AS·5AL-5RL chromosomes among 41 F5 plants from the cross CS ph1bph1b/MA5R, and aberrations permitted the location of the hairy peduncle gene (Hp) and marker XLFZ3473 to the region 5RL FL 0.78–1.0 with another nine markers locating to 5RL 0.0–0.78. The chromosome-specific markers and chromosome aberrations developed in this study will facilitate the introgression of rye chromatin into wheat.