Observations on the Fusion of Nuclei in Plants

During the life cycle of the flowering plant, there are normally three occasions when nuclei fuse: (1) when the two polar nuclei of the mature megaga-metophyte fuse; (2) when one of the two male nuclei fuses with the egg nucleus; (3) when the second male nucleus fuses with the combined polar nuclei. Observations on the actual fusion of the nuclei made with the light microscope are Recent observations made with the electron microscope and reported here have revealed the basic method of nuclear fusion in the cases mentioned above. MATERIALS AND METHODS Thc plant used was cotton (Gossypium hirsutum). The nuccllus was isolated at various times after the flower had opcned and had been hand pollinated. Thc tissuc was fixed in 2 per cent KMnO4 at 4°C for 4 hours or in 6 pcr cent glutaraldchydc (pH 6.8, 0.1 M phosphate buffer) at 4°C for 4 hours, washed in buffer, and placcd in 2 per cent OsO4 at 4°C for 12 hours. Thc dehydration was through an acetone scrics. To the 70 per cent acetone was added 1 per cent uranyl nitrate. All tissue was embedded in Epon and sectioned with a diamond knife. All observations were made with a Bendix Transcope RS-50-E-1. OBSERVATIONS The nuclei were observed to fuse in the following manner. The nuclei which will fuse migrate toward one another, but a layer of cytoplasm, including mitochondria, Golgi apparatus, sphero-somes, endoplasmic reticulum, and even small vacuoles, still separates them (Figs. 1 (A) 2). Next, at several points elements of the endoplasmic reticulum from the two nuclei fuse, making the outer nuclear membranes of the two nuclei continuous (Figs. 1 (B), 2 (arrows)). The membranes connecting the two nuclei apparently decrease in length and membranous bridges composed of the outer nuclear membranes are formed (Figs. 1 (C), 3). The inner nuclear membranes next come into contact and merge (Figs. 1 (D_), 3). Thus, first the outer and then the inner nuclear membranes fuse