Thinopyrum intermedium, a tertiary gene pool of wheat, harbors valuable genetic resources for wheat improvement. This study aimed to develop novel wheat germplasms by transferring elite chromosomes from Thinopyrum intermedium into wheat via distant hybridization, investigating their impacts on disease resistance and agronomic traits to establish a theoretical foundation for wheat breeding.

A wheat-Thinopyrum intermedium disomic addition line, CH71 (2n=44), was developed from the BC₁F₆ progeny of a cross between common wheat cultivar Yannong 999 and the partial amphidiploid TAI8047 (2n=58). Non-denaturing fluorescence in situ hybridization (ND-FISH) with oligonucleotide probes (Oligo-pSc119.2, Oligo-pTa535, Oligo-B11, Oligo-pDb12H) was employed for karyotype analysis. Synteny-based (Synt) Oligo-FISH painting and Th. intermedium-specific STS markers were utilized to identify the homologous group and validate the origin of the alien chromosome. Disease resistance to powdery mildew (race E09) and stripe rust (mixed races CYR32, CYR33, CYR34) was evaluated under artificial inoculation. Agronomic traits, including plant height, spike length, spikelet number, thousand-grain weight, grain length, and grain width, were systematically measured.

The wheat parent Yannong 999 exhibited a standard karyotype of 21 wheat chromosome pairs. TAI8047 contained 58 chromosomes (21 wheat pairs + 8 alien pairs), while CH71 harbored 44 chromosomes (21 wheat pairs + 1 alien pair, J^S-1). Synt-FISH and PCR amplification using 183 STS primers specific to Th. intermedium chromosome 7J^S confirmed J^S-1 as a 7J^S-derived chromosome. Twelve STS markers consistently amplified diagnostic bands in Th. intermedium, TAI8047, and CH71. CH71 displayed moderate resistance to powdery mildew and immunity to stripe rust, with genetic analysis indicating both resistances were conferred by the 7J^S chromosome. Compared to Yannong 999, CH71 exhibited significant increases in plant height (+24.0 cm), spike length (+3.44 cm), spikelet number (+1.6), and grain length (+0.68 mm), but reductions in thousand-grain weight (-6.78%) and grain width (-2.70%).

The novel disomic addition line CH71 (wheat-Th. intermedium 7J^S) demonstrates dual resistance to powdery mildew and stripe rust, serving as a valuable germplasm resource for disease-resistant breeding and the cloning of alien resistance genes. The 12 STS markers identified herein provide an efficient molecular tool for rapid tracking of the 7J^S chromosome in wheat backgrounds.