The wheat-alien small segment translocation (SAST) lines carrying the beneficial genes from wild species are useful genetic stocks for wheat improvement. In this study, to introduce the grain hardness-related genes of Haynaldia villosa (L.) Schur. into common wheat (Triticum aesitivum L.), the mature female gametes of whole-arm wheat--H. villosa translocation line T5VS·5DL was irradiated by 60CO-γ ray to develop SAST lines involving 5VS. Among the BC2F2 population, six homozygous SAST lines with different fragment sizes of 5VS were identified by GISH, and the exact fragment sizes were further defined using four 5VS-specific markers and four Ha gene-based markers. The results showed that five lines (NAU5VS-1 to NAU5VS-5) carried the softness gene Dina/Dinb of H. villosa, and that NAU5VS-5 had the smallest alien translocation segment, identified to be a 5VS-6AS·6AL terminal translocation. The translocation chromosome 5VS-6AS·6AL was proved to be stably inherited to the successive generations. In the BC3F2 generation, the individuals having the homozygous 5VS-6AS·6AL translocation chromosomes all showed soft grain texture, with an approximately 50% reduction in the SKCS hardness index compared with that of their backcrossing parent. Both the 5VS-6AS·6AL translocation line and the molecular markers developed in this study will be valuable in wheat breeding for soft grain quality improvement.