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Sinorhizobium meliloti forms a robust N₂-fixing root-nodule symbiosis with Medicago sativa. We are interested in identifying the minimal symbiotic genome of the model strain S. meliloti Rm1021. This gene set refers to the minimal genetic determinants required to form a robust N₂-fixing symbiosis. Many symbiotic genes are located on the 1,354 kb pSymA megaplasmid of S. meliloti Rm1021. We recently constructed a minimalized pSymA, minSymA2.1, that lacked over 90% of the pSymA genes. Relative to the wild-type, minSymA2.1 showed a reduction in M. sativa shoot biomass production and nodule size with an increase in total nodule number. Here we show that the addition of either the stachydrine (stc) or trigonelline (trc) catabolism genes from pSymA to minSymA2.1 restores nodule size and total nodule number to levels indistinguishable from the wild-type but does not restore reduced shoot biomass production. In the context of the complete Rm1021 genome, removing the stc genes reduced nodule size and increased total nodule number while removal of the trc genes alone had no apparent effect. Together, these observations implicate stachydrine catabolism as an important determinant of root nodule symbiosis between S. meliloti and M. sativa while trigonelline catabolism seems to contribute in a more conditional manner, in the context of the minimized genome. These findings highlight the minimal symbiotic genome as a tool for investigating the impact individual genetic determinants have in conferring an optimal symbiosis. Factors whose impact, in the context of a complete genome, may be hidden or dampened due to redundancies.