DAPI Staining and Fluorescence Microscopy

Anthers at stage 8 or 12 were collected and crushed with a fine syringe needle to release microspores or mature pollen grains in a drop of water on the slide. The newly released microspores were stained with 1 μg/mL of DAPI and observed immediately by fluorescence microscopy (Zeiss Imager M2).

For the B'α-YFP and B'β-YFP expression assays, ovules and anthers from the complemented plants were used. Ovules and male meiocytes, released from anthers at the meiosis stage, were stained with 1 μg/ml of DAPI and observed by confocal laser scanning microscopy (Zeiss LSM-710). Fluorescent signals were captured at 526–573 nm (emission) for YFP with 488-nm excitation and 420–490 nm (emission) for DAPI with 340-nm excitation.