In vitro manipulation of cucumber [Cucumis sativus L.] pollen and microspores: isolation procedures, viability tests, germination, maturation

The distribution of plastids at the time of microspore and pollen grain development in Gagea lutea (L.) Ker.-Gaw. was analyzed using electron microscopy. It was shown that plastids are not transmitted to the forming generative cell of this species during microspore division. At the vacuolate microspore stage, preceding division, the microspore nucleus takes an acentric position and the plastids gather at the opposite side of the cell. In the highly polarized microspore at prophase of mitosis, all plastids are aggregated at one side of the nucleus, whereas mitochondria are dispersed throughout the cytoplasm. Numerous profiles of endoplasmic reticulum (ER) are present between the clustered plastids. Some of the ER profiles are attached by their ends to the outer membrane of plastid envelopes and join the distant plastids. The outer membrane of the microspore plastids may form long and thin evaginations contacting with other plastids. Microtubules are visible in plastid aggregations occasionally. In dividing microspores, long ER cisterns surround the area of the mitotic spindle and separate it from the region containing plastids. There are no plastids in the young generative cell: all plastids remain clustered in the region of the microspore that now forms the vegetative cell of the bicellular pollen grain. Later the connections between plastids and ER cisterns gradually disappear and plastids disperse in the cytoplasm of the whole vegetative cell. The results of our study are not sufficient to define the mechanism causing selective aggregation of plastids at the vegetative pole of the Gagea microspore, nor to say whether the microtubular cytoskeleton plays a role. However, the participation of ER in these processes, at least in holding the special arrangement of microspore plastids, seems certain.