Utilization of Interspecific Heterosis for the Creation of Elite Germplasm: A Comprehensive Analysis of Medicago sativa × M. falcata F1 Hybrid Populations

In this study, 101 F1 hybrid lines were constructed using Medicago sativa L. ‘Xinjiang Daye’ and Medicago falcata L. ‘Hulunbeier’ as parents, and their agronomic and quality traits were systematically evaluated over two consecutive years. The results showed that the hybrid progeny exhibited more pronounced phenotypic variation in the second year. Specifically, the X4H4 combination demonstrated superior biomass accumulation, while the H4X4 combination showed notable advantages in quality indices. In the second year, the correlations among agronomic traits were clearly strengthened, with morphological traits closely associated with biomass; coordinated variation was also observed among quality traits, as crude protein content was negatively correlated with fiber-related indices. Based on principal component and heterosis analysis, the hybrid lines were classified into distinct advantage groups: Group 1 exhibited clear agronomic heterosis, while Groups 2 and 3 displayed distinct advantages in quality traits. Finally, 12 elite individuals were selected based on integrated KASP molecular marker profiling and comprehensive phenotypic evaluation. This study establishes a strategy for marker-assisted phenotypic evaluation in alfalfa breeding, providing a theoretical basis and germplasm resources for the development of high-yield and high-quality cultivars.