利用激光扫描共聚焦显微成像系统的正交切割成像确证GFP标记细菌在植物根内定殖  

Using orthogonal sectioning with a laser scanning confocal microscopic system to determine endophytic colonization of plant roots by GFP-tagged bacteria

作　　者：安千里[1] 杨学健[2] 冯利[1] 毛碧增[1] 匡柏健[2] 李久蒂[2] 

机构地区：[1]浙江大学生物技术研究所,浙江杭州310029 [2]中国科学院植物研究所,北京100093

出　　处：《电子显微学报》2010年第4期354-360,共7页Journal of Chinese Electron Microscopy Society

基　　金：浙江省自然科学基金资助资助项目(Y307143);浙江省科技厅钱江人才计划项目(2007R10037);浙江大学优秀青年教师资助计划项目(紫金计划)

摘　　要：用激光扫描共聚焦显微镜(LSCM)检测绿色荧光蛋白(GFP)标记的细菌常用来确证植物内生细菌的内生特性。本文通过用GFP标记的内生细菌侵染植物根,用LSCM分别检测植物的根段或根的徒手横切片,显示LSCM系统的无损伤光学切片和正交切割成像适用于确证GFP标记细菌在植物根内定殖,显示徒手切片对揭示细菌定殖模式及其入侵途径的不良影响,并依此更新LSCM检测GFP标记细菌在植物根内定殖的方法。Laser scanning confocal microscopy（LSCM） to detect gfp-tagged bacteria in plants is widely used to determine bacterial endophytic nature.LSCM detected colonization in roots of sugarcane micropropagated plantlets and maize seedlings by a gfp-tagged endophytic nitrogen-fixing bacterial strain,Klebsiella oxytoca SATC3.LSCM orthogonal sectioning through intact root pieces demonstrated bacterial colonization in root cortical and stele tissues and in root hair cells.LSCM orthogonal sectioning could demonstrate bacterial endophytic colonization unequivocally but with a low resolution.LSCM optical sectioning through hand-cut root transverse sections revealed bacterial colonization in intercellular spaces in primary and lateral root cortices and in stele cells including xylem vessels and their adjacent cells.However,hand-cut sectioning cause artifacts of bacterial translocation and root breakage.Therefore,LSCM orthogonal sectioning of intact roots should be used to demonstrate bacterial endophytic colonization and to find the locations for further microscopic analysis of the exact bacterial localization and of interactions between the endophytic bacteria and plant cells.

关 键 词：内生细菌 绿色荧光蛋白 激光扫描共聚焦显微镜 正交切割 

分 类 号：Q938[生物学—微生物学] Q-336