唐菖蒲花粉原生质体及其萌发花粉管的超微结构研究

AN ULTRASTRUCTURAL STUDY ON POLLEN PROTOPLASTS AND POLLEN TUBES GERMINATED FROM THEM IN GLADIOLUS GANDAVENSIS

用酶法从唐菖蒲(Gladiolus gandavensis)成熟花粉粒中分离出大量原生质体。在培养条件下,诱导了细胞壁再生和花粉管萌发,萌发率达47.7％。电镜观察表明:花粉原生质体呈圆球形,仅为质膜包围,内部超微结构完整无损。再生壁与原来的花粉壁不同,呈多层次的纤丝结构,有类似萌发沟处的局部增厚。许多小泡参与了壁的合成。线粒体更为丰富,质体中出现少量淀粉,游离核糖体与油滴增多。花粉原生质体由萌发沟处长出花粉管。培养18小时,营养核和生殖细胞移入花粉管,其周围质体丰富。高尔基体分布在花粉管边缘。有小泡溢出质膜外,参加花粉管壁的合成。

Large quantities of protoplasts were isolated enzymatically from the mature pollen grains in Gladiolus gandavensis. Regeneration of cell wall and germination of pollen tubes were per- formed during culture of purified pollen protoplasts in K_3 medium supplemented with 32% su- crose, 0.1 mg/l 2,4--D, 1 mg/l NAA and 0.2 mg/l 6--BA, with a germination rate up to 47.7%. The materials were fixed gently with gradually increasing concentration of glutaraldehyde, fol- lowed by osmium, then preembedded in a thin layer of agar and surveyed under an inverted microscope so as to select desired specimens for subsequent procedure. Small agar blocks con- taining specimens were dehydrated through ethanal-propylene oxide series, embedded in Aral- dite and ultratomed. Electron microscopic observations show that the pollen protoplasts are surrounded by a smooth plasma membrane and with ultrastructurally intact cytoplasm, a vege- tative nucleus and a generative cell. After 8h of culture, wall regeneration commences resulting in a multilayered, fibrillar wall structure which is different from the intine. No exine is formed. Numerous vesicles participate actively in the wall formation. The wall is uneven in thi- ckness around its periphery; a thickened area somewhat resembling to germ furrow is formed, from which pollen tube emerges. The tubes contain abundant plastids, mitochondria and dictyo- somes. Vesicles are released out of the plasma membrane and involved in tube wall formation. After 18h of culture, the vegetative nucleus and generative cell have migrated into the tube. Te- chnical points of preparing pollen protoplast specimens for ultastructural studies and the featu- res of wall regeneration in pollen protoplast culture are discussed.