Abstract
Pollen development in angiosperms starts in anthers within buds formed by conversion from vegetative growth to reproductive growth, and results in double fertilization at which one sperm cell fuses to an egg cell and the other sperm cell fuses to a central cell including two polar nuclei. During this process, there are drastic cellular changes such as production of haploid cells by meiosis in pollen mother cells, differentiation of the generative cell and the vegetative cell by an unequal (asymmetric) cell division in microspores and growth of pollen tubes. However, it is generally difficult to observe the inner structure under an ordinary light microscope because the pollen mother cells possess callosic cell wall, and microspores and pollen grains further have a thick exine consisting of insoluble sporopollenin. Therefore, the observation has usually been conducted on sectioned samples as in the case with electron microscopy. In order to make the observation easy, we tried to isolate protoplasts free from cell wall and exine in Lilium longiflorum. As a result, pollen mother cell protoplasts, microspore protoplasts, pollen protoplasts and generative cell protoplasts could be obtained in large quantities. On the other hand, biochemical research suggested several specific proteins appeared during pollen development. Using specific antibodies raised against the specific antigens, immunofluorescence studies on isolated protoplasts first revealed dynamics of the specific proteins during pollen development.
