AbstractFollowing assimilation of14CO2by leaves ofStachys sieboldii,14C‐stachyose is translocated into the tubers. Stachyose is accumulated and stored in the vacuoles of the pith parenchyma. Protoplasts and vacuoles were isolated and the uptake of sugars was examined. Uptake of sucrose and sucrosyl oligosaccharides of the raffinose family by protoplasts was very low compared to glucose. Transport parameters for glucose indicated a carrier mediated transport in the lower concentration range which was superimposed by diffusion at higher concentrations (>10 mM). The very low sugar uptake by protoplasts and the sparse enzyme activities of stachyose synthase in the storage parenchyma as well as acid invertase and α‐galactosidase in the cell walls indicated symplastic unloading of stachyose in the tubers. Experiments on14C‐stachyose uptake by isolated vacuoles confirmed previous observations by Keller (1992).Isolated vacuoles exhibited ATP and PP hydrolysis and were capable of generating a proton gradient across the tonoplast by a V‐type H+‐ATPase and H+‐PPase. This was demonstrated by fluorescence quenching of quinacrine. Fluorescence could be restored by the addition of gramicidin and partly recovered by the addition of stachyose; mannitol, sorbitol and glucose had no effect. Fluorescence recovery depended on the concentration of stachyose and revealed saturation kinetics (Km= 28 mM). Comparable results have been obtained with tonoplast vesicles by Greutert and Keller (1993).Experimental data presented here provide circumstantial evidence for symplastic unloading of stachyose in the tubers ofStachys sieboldiiand demonstrate that the stachyose concentration in the cytoplasm of storage parenchyma cells is kept low by active stachyose transport into the vacuoles. The results suggest a stachyose/H+‐a