Determination of H-ATPase Activity in Guard Cell Protoplasts through H-pumping Measurement and H-ATPase Quantification.

The opening of stomata in plants in response to blue light is driven by the plasma membrane H-ATPase in guard cells. To evaluate the activation of the H-ATPase , we can use H-pumping by guard cells in response to blue light and fusicoccin. To do this, it is required to prepare a large amount of guard cell protoplasts and measure H-pumping in the protoplasts. It is also necessary to determine the protein amount of H-ATPase. In this protocol, we describe the procedures required for these preparations and measurements.