Characterization of a Natural Accession of Elymus sibiricus with In Situ Hybridization and Agronomic Evaluation

<i>Elymus sibiricus</i>, valued for its perennial nature, broad adaptability, strong cold tolerance, and high economic value in forage production, plays a crucial role in combating grassland degradation, desertification, and salinization. Using morphological and cytogenetic methods, this study evaluated the cold tolerance, post-harvest regeneration capacity, and perennial characteristics of the <i>E. sibiricus</i> accession 20HSC-Z9 in the Harbin region of China from 2020 to 2023. This accession exhibited a germination rate of over 90% and a 100% green-up rate, with purple coleoptiles indicating its strong cold tolerance. Over the three growing seasons, 20HSC-Z9 maintained stable green-up and regeneration rates, confirming its perennial nature. Morphologically, 20HSC-Z9 had an average tiller count ranging from 56 to 74, similar to that of the control accession 20HSC-ES, and its plant height was significantly lower than that of 20HSC-IWG. Furthermore, 20HSC-Z9 produced over 100 grains per spike, with a seed setting rate exceeding 90%, and a thousand-grain weight comparable to that of 20HSC-IWG. The grain protein content of 20HSC-Z9 reached a maximum of 21.19%, greater than that of the control accessions (15.6% and 18.5%). Chromosome composition analysis, using sequential multicolor genomic <i>in situ</i> hybridization and multicolor fluorescence <i>in situ</i> hybridization, confirmed the StStHH genomic constitution of 20HSC-Z9 and revealed translocations between the St and H subgenome chromosomes. These results suggest that 20HSC-Z9 has significant potential as a new perennial forage grass germplasm for cold regions, suitable for further domestication and breeding efforts.