Alfalfa (Medicago sativa L.), an important perennial legume forage crop with high nutritional value and forage yield, is widely used in animal husbandry. However, it is very sensitive to aluminum, which severely limits its growth in acidic soils. In this study, we analyzed the genotype variation of each agronomic trait in 44 alfalfa varieties in two acidic soil environments. Then, analysis of variance (ANOVA) of the variance components was performed using the Residual Maximum Likelihood (REML). The best linear unbiased predictor analysis was used to obtain the mean trait of each variety, and the mean values were used to construct the mean matrix of varieties × traits and interaction analysis of varieties × years. The results showed that there was significant (P < 0.05) genotypic variation for each trait of the 44 varieties and the genetic diversity was abundant. The average repeatability (R value) of interannual plant height (PH), stem thickness (ST), number of branches (NS), fresh weight (FW), total fresh weight (TFW), and total dry weight (TDW) was high (0.21–0.34), whereas the genetics were relatively stable. PH, NS, FW, TFW, and dry weight (DW) were positively correlated (P < 0.01) with TDW. Six alfalfa varieties (Algonquin, Xinjiang daye, Trifecta, Vernal, WL354HQ, and Boja) with excellent TDW and TFW were identified in different years, environmental regions, and climatic altitudes. Our research results can provide suggestions and critical information regarding the future improvement and development of new alfalfa strains and varieties that are resistant to acidic soil conditions.