V-ATPase, PPase activity assays and vacuolar pH measurements in OsVHA-A RNAi transgenic lines.

(A) Vacuolar H+-ATPase activity and (B) V-PPase activity were determined in wild type (WT) and three OsVHA-A RNA interference lines (OsV-5, OsV-11, and OsV-18). (C) The images showing emission intensities of vacuoles from epidermal root cells loaded with BCECF AM. Results shown are representative. Scale bars  = 100 μM. (D) The vacuolar pH values calculated from (C). (E) V-ATPase proton-pumping measured by the quenching of ACMA fluorescence. Ten micrograms of tonoplast vesicles were applied to detect fluorescence density. Each bar represents three replications. Asterisks (*) indicate significant differences from WT at P<0.05.