利用激光共聚焦扫描显微镜对溃疡病菌侵染柑橘叶片的实时观察  

Real Time Observation of the Infection of Xanthomonas citri subsp.citri in Citrus Leaf by Using Confocal Laser Scanning Microscopy

作　　者：赵亚[1] 马先锋[1] 肖翠[1] 刘利平[1] 李娜 邓子牛[1] 

机构地区：[1]湖南农业大学园艺园林学院,国家柑橘改良中心长沙分中心,湖南长沙410128

出　　处：《激光生物学报》2017年第2期143-150,共8页Acta Laser Biology Sinica

基　　金：国家自然科学基金(31572111)

摘　　要：柑橘溃疡病对柑橘产业造成了巨大损失,而研究柑橘与溃疡病菌的互作关系以及柑橘的感病和抗病性均需要观察溃疡病菌在柑橘寄主中的侵染和定殖过程。激光共聚焦扫描显微镜不仅可以观察活细胞,活组织的动态代谢过程,而且可以获得三维图像,对于病原菌在柑橘植物组织内的繁殖和致病机制研究具有重要意义。但是,选择适宜的植物材料和制片方法对激光共聚焦扫描显微镜的观察效果影响很大。本文对激光共聚焦扫描显微镜所观察的材料在其处理和观察方法上加以改进,获得了质量更好的图片和实验结果,也使得实验更为方便快捷。激光共聚焦扫描显微观察还在瞬时表达分析中得到应用,提高了柑橘瞬时表达分析的效果。通过将切片和压片相结合观察到溃疡病菌在不同时间点对柑橘叶片的侵染情况,而通过3D建模能观察到柑橘叶片不同组织层面中的病菌数量和病菌位置,为研究溃疡病菌在叶片中的定殖方式和入侵数量提供了前期基础。Citrus canker caused a huge losses to the citrus industry in our country, therefore, the observation of infec-tion and colonization of the pathogen in citrus is indispensable to the research of interaction between citrus and Xan-thomonas citri subsp. citri （ Xcc） as well as the susceptibility and resistance of citrus. Confocal laser scanning microsco-py can as well observe the dynamic metabolism of living cells and living tissues, it can obtain the 3-D image as well, it is of great significance to the study of the propagation and pathogenic mechanism of pathogenic bacteria in citrus tissues. However, the selection of appropriate tissue materials and production methods have great influence on the observation effect of observations. In this paper, the improvement of material handling and observing methods of confocal laser scan-ning microscopy was made to obtain better quality images and experimental results; By comparing the old and new two experimental methods, more convenient and efficient experimental method was got ; The effect of transient expression a-nalysis of citrus was improved by the application of confocal laser scanning microscopy . The process of the infection of citrus leaf by Xcc at different time points was preliminaryly studied by combining the section and tabletting methods and the number of bacteria and germs in different levels of organization are observed by 3D model, which provide the basis for the study of bacterial canker in leaf colonization and invasion quantity.

关 键 词：激光共聚焦显微镜 柑橘溃疡病菌 入侵 

分 类 号：S182[农业科学—农业基础科学]