Selection of High-Yield Varieties (Lines) and Analysis on Molecular Regulation Mechanism About Yield Formation of Seeds in Alfalfa

The goal of this study was to elucidate the genetic and molecular regulatory mechanisms underlying agronomic traits in elite alfalfa (Medicago sativa L.). Through the analysis of 44 varieties and lines, we measured 19 yield-related traits and performed transcriptome sequencing to investigate the factors driving yield variation. The results indicated extensive variation in agronomic traits among the tested accessions, with the coefficients of variation (CVs) ranging from 7.85% to 42.66%, suggesting substantial potential for genetic improvement. Correlation analysis revealed that seed yield was significantly and positively correlated with the number of reproductive branches and inflorescences at maturity, whereas early vegetative growth was negatively correlated with 100-seed weight. The 44 accessions were categorized into three clusters: Cluster II (the largest group) exhibited balanced traits; Cluster I showed vigorous early growth but low pod yield; and Cluster III was characterized by the highest pod and branch numbers. Principal Component Analysis (PCA) explained 65.88% of the total variation (first six components), identifying GNS31 and GNS43 as the superior and inferior genotypes, respectively. Furthermore, transcriptome profiling detected the highest number of differentially expressed genes (10,089 DEGs) in pod tissues, with 66% being upregulated. Functional enrichment analyses (GO and KEGG) highlighted that varietal differences were primarily enriched in secondary metabolism, lipid metabolism, and plant hormone signal transduction pathways. Notably, within the auxin pathway, the SAUR and GH3 families displayed significant tissue-specific expression in pods.