植物内生菌透射电镜负染标本制备方法的对比研究  

Comparative Study on the Preparation Methods of Transmission Electron Microscope Negative Staining Specimens for Plant Endophyte

作　　者：黄远洁[1] 李卫东[1] 莫肖敏[1] HUANG Yuanjie LI Weidong MO Xiaomin(Electron Microscope Laboratory of the Medical and Scientific Research Center, Guangxi Medical University, Nanning Guangxi 530021, China)

机构地区：[1]广西医科大学医学科学实验中心电镜室,广西南宁530021

出　　处：《广西师范大学学报（自然科学版）》2016年第3期127-130,共4页Journal of Guangxi Normal University:Natural Science Edition

基　　金：国家自然科学基金资助项目(81560608);广西自然科学基金资助项目(2015GXNSFAA139123);广西医科大学青年科学基金资助项目(GXMUYSF2014022)

摘　　要：通过比较4种不同方法处理的植物内生菌负染标本的透射电镜检测结果,探讨植物内生菌透射电镜负染标本的理想制备方法。采用直接刮取法、双蒸水冲洗法、双蒸水冲洗离心重悬法和双蒸水冲洗静置法制备菌悬液,应用10g/L磷钨酸漂浮法负染,透射电子显微镜检测,根据菌的形态特征及菌毛、鞭毛形状数量等指标判断取材及处理方法的各自优缺点。结果显示:直接刮取法的样品原始状态保存较好,但杂质多,背景重;双蒸水冲洗的样品背景较干净;双蒸水冲洗离心重悬法和双蒸水冲洗静置法的样品视野清晰、分布均匀,背景干净、杂质少,但前者菌的形态结构较易受影响。可见,透射电镜是植物内生菌检测的重要工具,需根据实际情况选用样品制备方法。在严格控制静置时长的前提下,双蒸水冲洗静置法制备菌悬液后应用负染技术可获得较理想的检测效果。Abstract： The results of transmission electron microscope tests of the negative staining specimens of plant endophyte were comparatively studied to explore the ideal preparation methods of transmission microscope negative staining specimens of plant endophyte. Firstly, four methods were used to prepare endophyte suspension, such as direct scraping, double distilled water flushing, double distilled water flushing followed by centrifugal separation and re- suspension and double distilled water flushing followed by static placing. Then, 10 g/L phosphotungstic acid was applied for floating negative staining and the transmission electron microscope test was carried out. And finally, the morphological characteristics, the shape and the quantity of the pilus and the flagellum of the endophyte were used as indicators to assess the advantages and disadvantages of these extracting and processing methods. Results： （1） The original state of the direct scraping sample is better, hut the background is heavy, and the surface of the cell is affected easily by impurities. （2） The sample background of the double distilled water flushing is better. （3） The sample of the double distilled water flushing followed by centrifugal separation and re--suspension, and the sample of the double distilled water flushing followed by static placing have a clear vision, uniform distribution, clean background and less impurity under the transmission electron microscope. But the morphology structure of endouphyte of the former one was more easily affected. Transmission electron microscopy is an important tool for endophyte research. However, it is necessary to choose sample processing methods based on the actual situation. Under strict control of the static time, double distilled water flushing followed by static placing with the negative staining technique can achieve better test results.

关 键 词：植物内生菌 透射电镜 负染 

分 类 号：Q93-33[生物学—微生物学]