Cytogenetic identification and molecular marker development of a novel wheat-tetraploid Thinopyrum elongatum small fragment translocation line with stripe rust resistance gene Yr4EL

Tetraploid Thinopyrum elongatum (2n = 4x = 28, EEEE) is a valuable genetic resource for wheat improvement, harboring genes conferring resistance to multiple diseases, including stripe rust, powdery mildew, and Fusarium head blight. In our previous study, we developed the wheat-tetraploid Thinopyrum elongatum translocation line T4DS·4DL-4EL, which carries a 49.59 Mb terminal segment of chromosome 4EL containing the Yr4EL stripe rust resistance locus. In this study, the ph1b mutant was used to promote homoeologous recombination and reduce the size of the 4EL introgression. Four novel small-fragment T4DS·4DL-4EL translocation lines carrying terminal 4EL segments of 13.22 Mb, 1.99 Mb, 1.42 Mb, and 1.08 Mb were identified and characterized using GISH, FISH, and molecular markers. Stripe rust response evaluation showed that only T4DS·4DL-4EL#3 exhibited all-stage resistance to stripe rust, and genetic analysis demonstrated complete association between the 13.22 Mb 4EL introgression and resistance in the F₂ population. Two Kompetitive Allele-Specific PCR markers were developed to distinguish plants carrying the 13.22 Mb 4EL introgression and showed complete concordance with the resistance phenotype in the F₂ population. By integrating the breakpoint information from these translocation lines with our previously reported genetic mapping results, the candidate interval of Yr4EL was narrowed to approximately 0.8 Mb (610.2–611.0 Mb), flanked by the molecular markers TTE4E-7767 and TS4E-1915. Three specific Sequence-Tagged Site markers co-segregating with this 0.8 Mb interval were also developed. Collectively, the small fragment translocation lines developed in this study provide new germplasm resources for gene cloning and breeding utilization.