小麦条锈菌胞质游离钙离子动态检测方法的建立  

A detection method for intracellular calcium ion of Puccinia striiformis f. sp. tritici

作　　者：张洪[1] 丁可[1] 裴国亮[1] 郭军[1] 康振生[1] 

机构地区：[1]西北农林科技大学植物保护学院陕西省农业分子生物学重点实验室,杨凌712100

出　　处：《菌物学报》2010年第1期64-67,共4页Mycosystema

基　　金：国家自然科学基金(No.30671350);教育部科学技术研究重点项目(No.107104);教育部长江学者和创新团队发展计划项目(No.IRT0558);高等学校学科创新引智计划资助项目(No.B07049)

摘　　要：胞质游离钙离子变化与植物病原真菌侵染寄主的动态过程具有重要的关联性。本研究以侵染小麦叶片的条锈菌31号生理小种(CYR31)为材料,以孵育法将Ca2+荧光探针Fluo-3-AM载入到小麦条锈菌细胞中,并结合激光共聚焦扫描显微技术,建立了测定侵染过程中条锈菌胞质游离Ca2+分布的试验方法。结果表明,采用10μmol/L Fluo-3-AM装载顺次进行低温4℃孵育1h,25℃孵育1h,可获得较为理想的条锈菌胞质游离Ca2+染色结果。该方法可用于检测不同侵染阶段的小麦条锈菌细胞质游离钙离子的分布变化,为进一步研究锈菌胞内钙离子动态与侵染寄主的关联性提供了技术支撑。Calcium dynamics during the interactions between Puccinia striiformis f. sp. tritici （Pst） and wheat is essential for understanding fungal infection process associated with intracellular free calcium in Pst. In this study the leaves of wheat cultivar ＇Suwon 11＇ infected by urediospores of Pst CYR31 race were used to optimize calcium labeling method. Using fluorescence dye Fluo-3-AM （10μmol/L） to incubate with Pst cell, we successfully established an optimal method for detecting intracellular free calcium in the cells of Pst with a confocal laser scanning microscopy. The methodological procedure included to first incubate the cells with staining dye at 4℃ for 60 minutes and at 25℃ for 60 minutes. The method established in this study could be used to detect intracellular free calcium in Pst and further investigate its association with fungal pathogenesis against wheat host.

关 键 词：小麦条锈菌 胞质游离钙离子 动态检测方法 荧光指示剂 

分 类 号：S435.121[农业科学—农业昆虫与害虫防治]