Comparison Of Ultrastructure And Micro-quantitative Analysis Of Multi-spectral In Development Of Pollen In Hybrid Rice

Male sterility is the biological basis of heterosis utilization in crop breeding, especially in rice (Oryza sativa L.). Therefore, research on rice male sterility is a fundamental field of plant genetics and breeding. In this paper, we compared the developments of sterile anthers and the features of pollen abortion between Zhenshan 97 and YunNan Purple rice by electron microscope. In addition, for the first time, we investigated polysaccharides produced in anther development in both four cytoplasmic male sterility lines including Honglian, Maxie,Zhenshan 97 and zidao and their maintainer lines by the multi-spectralimaging and micro-quantitative analysis of big molecular substances. The results are mainly as follows.1. YunNan Purple rice was a new cytoplasmic male sterility line belonged to non-pollen type abortion. The vacuolated cytoplasm took place at the early stage of microspore development, showing a trend of abortion. Additionally, the abortion occurs earlier compared with that of CMS-Zhenshan 97 rice. At the stage of pollen abortion, tapetum cells are active, represented by thick cytoplasm and clear boundary. Our data suggested the abnormal development of tapetum cells was probably one of the reasons led to the cytoplasmic male sterility of YunNan Purple rice.2. In the maintainer lines, multi-spectral imaging of polysaccharides showed a V-shaped curve, with the lowest value appear in the wavelength range from 550 nm to 570 nm. When the anther of male sterility line became abortive, curves of polysaccharides showed a sudden decrease at wavelength of 420 nm. We found that this decrease was closely related to the time of pollen abortion of rice, which in turn can be applied to determine the abortion period of rice.3. Observations on anther polysaccharide dynamics change of rice maintainer lines revealed that pollen mother cells were stained strongly by Periodic Acid Schiff (PAS), with small but abundant grains during the process of pollen mother cell meiosis. At the same time, the amount of polysaccharides in the cells increased slightly. In contrast, polysaccharide grains of uninucleate-stage pollen were bigger, more distinctive and there was a decrease in the amount of polysaccharides. At the late stage of uninuleate cell, the amount of polysaccharides reached its lowest. At the binucleate-stage pollen, polysaccharide grains became diffused, resulting in an increase of polysaccharides. At maturity, fertile pollen developed and produced ample starch grains, together with a highest value of polysaccharides (starch) in pollen cells.4. Cytoplasmic male sterility line differed from its maintainer lines not only by multi-spectral curves but by the amount of polysaccharides. At both tetrad and pollen mother cell stages, the amount of polysaccharides in pollen cells of male sterility line was slightly higher than that of its maintainer lines. Experiments further revealed that there was no significantly difference between them. At the early microspore stage, the amount of polysaccharides of CMS Honglian, Maxie and Zhenshan 97 was similar to that of its maintainer lines, but the amount of polysaccharides of CMS Zidao was significantly higher than that of its maintainer line. At late microspore and bicellular pollen stage, the amount of polysaccharides of CMS Honglian and Maxie was significantly lower than that of its maintainer lines. At the abortive stage, the amount of polysaccharides in abortive pollens was significantly lower than that of mature pollens.In addition, we carried out an experiment on the development of SSR marker of nuclear genome of the two representative species of the genus Sparganium, namely S. glomeratum and S. simplex. By FIASCO, we developed 54 pairs of SSR marker of nuclear genome in the species Sparganium glomaratum. Among them,13 pairs of them are effective but only 9 pairs belonged to polymorphic Simple Sequence Repeat (SSR) markers. Furthermore, cross-species amplification in five individuals of S. simplex showed 7 of 9-pair markers was successful to the amplification. At last, nine pair SSR markers was applied to test for the genetic variation between 45 individuals of Sparganium glomaratum from different regions of the northeast China. The results revealed that the samples from the same gathering localities clustered into a group. These findings will contribute to the understanding of the genetic variation and evolution and distributional pattern of the genus Sparganium.