Screening of environmental stimuli for the positive regulation of stomatal aperture in centipedegrass

Grasslands, the largest carbon pool in China, possess enormous potential for carbon sequestration. Increasing the stomatal aperture to increase the CO₂ absorption capacity is a potential method to improve plant photosynthetic efficiency and ultimately enhance the carbon sequestration capacity of grass plants. Research on stomatal aperture regulation has focused mostly on Arabidopsis or crops, while research on grass plants in these areas is scarce, which seriously restricts the implementation of this grassland carbon sequestration strategy. Here, a widely used ecological grass, centipedegrass, was used as the experimental material. First, a convenient method for observing the stomatal aperture was developed. The leaves were floated in a potassium ion-containing open solution (67 mM KCl, pH 6.0) with the adaxial surface rather than the abaxial surface in contact with the solution and were cultivated under light for 1.5 h. Then, nail polish was applied on the adaxial surface, and a large number of open stomata were imprinted. Second, with the help of this improved method, the concentration‒response characteristics of the stomatal aperture to eleven environmental stimuli were tested. The stomatal aperture is dependent on these environmental stimuli in a concentration-dependent manner. The addition of 100 μM brassinolide led to the maximal stomatal aperture. This study provided a technical basis for manipulating stomatal opening to increase the carbon sequestration capacity of centipedegrass.