Cobalt (Co) contamination in soils potentially affects human health through the food chain. Although rice (Oryza sativa) as a staple food is a major dietary source of human Co intake, it is poorly understood how Co is taken up by the roots and accumulated in rice grain. In this study, we physiologically characterized Co accumulation and identified the transporter for Co uptake in rice. A dose-dependent experiment showed that Co mainly accumulated in rice roots. Further analysis with LA-ICP-MS showed Co deposited in most tissue of the roots, including exodermis, endodermis and stele region. Co accumulation analysis using mutants defective in divalent cation uptake showed that Co uptake in rice is mediated by the Mn/Cd/Pb transporter OsNramp5, rather than OsIRT1 for Fe and OsZIP9 for Zn. Knockout of OsNramp5 enhanced tolerance to Co toxicity. Heterologous expression of OsNramp5 showed transport activity for Co in Saccharomyces cerevisiae. Co uptake was inhibited by either Mn or Cd supply. At the reproductive stage, the Co concentration in the straw and grains of the OsNramp5 knockout lines was decreased by 41%-48% and 28%-36%, respectively, compared with that of the wild-type rice. The expression level of OsNramp5 in the roots was not affected by Co. Taken together, our results indicate that OsNramp5 is a major transporter for Co uptake in rice, which ultimately mediates Co accumulation in the grains.