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The success of a Rhizobium sp. inoculant depends on a product with a high number of live bacteria that can nodulate a leguminous plant. Culture media and oxygen transfer were studied to improve the production of R. phaseoli in chemically defined and undefined media for scale-up from shake flasks to a pneumatic bioreactor. The effects of carbon and phosphate on R. phaseoli growth were evaluated, and the specific volumetric oxygen transfer coefficient (KLa) was determined in shake flasks and scale-up to bioreactors. Six-fold more viable biomass was obtained by increasing the phosphate concentration by 7.5-fold when using glucose. However, this increase was not observed when succinate was used. In the undefined medium, glucose induced a higher production of R. phaseoli in comparison with other carbon sources. The KLa of 6–11 h-1 produced a five-fold growth of biomass in shake-flask cultures. This KLa was used to produce a R. phaseoli-based inoculant in a bioreactor. The formation of nitrogen-fixing nodules was confirmed in bean plants (Phaseolus vulgaris). The improvement of culture media with a suitable oxygen transfer demonstrates that R. phaseoli can be used as an inoculant that exerts beneficial effects on the initial growth of bean plants.