Microscopic Analysis of Spikelet and Anther

The plants were imaged with a Nikon D7000 digital camera (Nikon, Japan) and an Olympus SZX10 dissecting microscope (Olympus, Japan). Images of spikelets and anthers were captured with an Olympus BX51 fluorescence microscope (Olympus, Japan). Pollen viability was assessed using Alexander’s solution for staining wild-type and mutant anthers. By the use of semi-thin sections along with SEM (JEOL, Japan) and TEM (Hitachi, Japan) observations, the spikelets and anthers of wild type and osms188 mutant plants were classified as belonging to different stages to avoid experimental deviation. The embedding and observation procedures were performed as described in a previous study (Lou et al. 2014 (link)).