BACKGROUND: Flowering date in annual plants, and perception of symbiotic nitrogen-fixing rhizobium in legumes, are strongly selected traits. We asked if natural selection can be detected at key genes of the flowering and rhizobia recognition pathways in the model legume Medicago truncatula. RESULTS: We examined nucleotide variation among 57 accessions from natural populations in 53 gene fragments: 11 genes involved in flowering, 5 genes involved in nitrogen-fixing bacteria recognition, and 37 genes used as control loci. We detected 1757 polymorphic sites yielding an average nucleotide diversity of 0.003 per site. We found that accessions were structured in two groups consistent with geographical origins and that nucleotide diversity was not consistent with a neutral, constant-sized population but rather suggests recent expansion yielding more rare alleles than expected. Using coalescent simulations, we detected strong departure from selective neutrality in flowering genes within each group and showed that genetic differentiation between groups at several nodulation and flowering genes has probably been driven by recent selection. CONCLUSION: We quantify the intensity of purifying selection in M. truncatula and show that putative footprints of natural selection can be detected at different time scales in both flowering and symbiotic pathways.