水稻籽粒胚乳的石蜡切片方法改良及其结构发育观察

Improvement of Paraffin Section Methods and Structural Observation to Endosperm Development of Rice Kernel

以杂交水稻保持系‘岗46B’授粉后不同天数的籽粒为材料，利用PAS反应原理，通过对传统石蜡切片技术中材料固定、染色、脱水、浸蜡等环节的改进，即采用FAA和FPA相结合方式固定材料，先用热配法席夫（Schiff）试剂进行整染，再以稀释和氨水蓝化后的埃利希氏（Ehrlich）苏木精复染，中间以70％、83％、95％质量分数梯度乙醇进行脱水和复水处理，渐进式浸蜡与把握时间等实验环节，对水稻胚乳结构发育进行观察研究，结果授粉后第2～10天籽粒胚乳的石蜡切片完整，且染色分明，结构显示完整清晰。观察发现，随着授粉后天数推移，胚乳内部逐渐由游离胚乳核期向细胞化过渡，细胞层数增加并趋向充满子房，淀粉粒开始累积，胚乳细胞进一步增大。授粉后第5～10天是胚乳细胞淀粉粒从开始到大量累积的重要时期，同时胚乳细胞向无核化发展，不同部位胚乳细胞和淀粉粒的排列分布也逐渐表现出一定的差异。表明改良后的石蜡切片方法适合于授粉后第0～10天的水稻胚乳结构发育研究。

According to the principle of periodic acid-Schiff （PAS） reaction,we made some improvements about the rice endosperm structure of paraffin section. Using this improved method, we achieved the paraffin section of rice endosperm from the first to tenth day after pollination. In contrast to traditional method,we improved several steps, including fixation, staining, dehydrating, wax impregnation and so on. Fixation with FAA and FPA, whole staining with Schiff＇ s reagent, re-staining with Ehrlich hematoxylin diluted and blued by ammonia, dehydrating and saturating with grads alcohol of 70%, 83%, 95%, gradually immersing wax,all these measures achieved slicing up to the rice kernels of ＇Gang 46B＇ from the second to tenth day after pollination. The structure and staining of the tenth days＇ endosperm is clear and complete, which could not be realized by traditional method. We found that the inner endosperm gradually developed from the stage of free endosperm nuclear to cellularization. The cell layers in endosperm increase and fulfill the ovary. Starch granules start accumulation and gradually increase more. The volume of endosperm cell further increases. The fifth to tenth days after pollination are very important for starch granule accumulation in endosperm cell. At the same time,endosperm cell go into denucleation developing stage. The distributions of endosperm cell and starch granule are different as in different position of endosperm. The above results show that our improved method is suitable to be used in the study on endosperm development of rice kernel.