Department of Plant Anatomy, Institute of Biology, Eötvös Loránd University, 1117, Budapest, Pázmány Péter sétány 1/c, Hungary.
Mycorrhiza. 2014 May;24(4):259-66. doi: 10.1007/s00572-013-0533-8. Epub 2013 Nov 13.
In planta detection of mutualistic, endophytic, and pathogenic fungi commonly colonizing roots and other plant organs is not a routine task. We aimed to use fluorescence in situ hybridization (FISH) for simultaneous specific detection of different fungi colonizing the same tissue. We have adapted ribosomal RNA (rRNA) FISH for visualization of common mycorrhizal (arbuscular- and ectomycorrhiza) and endophytic fungi within roots of different plant species. Beside general probes, we designed and used specific ones hybridizing to the large subunit of rRNA with fluorescent dyes chosen to avoid or reduce the interference with the autofluorescence of plant tissues. We report here an optimized efficient protocol of rRNA FISH and the use of both epifluorescence and confocal laser scanning microscopy for simultaneous specific differential detection of those fungi colonizing the same root. The method could be applied for the characterization of other plant-fungal interactions, too. In planta FISH with specific probes labeled with appropriate fluorescent dyes could be used not only in basic research but to detect plant colonizing pathogenic fungi in their latent life-period.
在植物体内同时检测通常定植于根和其他植物器官的共生、内生和病原真菌并非常规任务。我们旨在使用荧光原位杂交（FISH）技术同时特异性检测定植于同一组织的不同真菌。我们已经针对不同植物物种根内的常见共生菌（丛枝菌根和外生菌根）和内生真菌，对核糖体 RNA（rRNA）FISH 进行了适应性改造。除了通用探针外，我们还设计并使用了特异性探针，这些探针与 rRNA 的大亚基杂交，并选择了荧光染料，以避免或减少对植物组织自发荧光的干扰。我们在此报告了 rRNA FISH 的优化有效方案，并报告了使用荧光显微镜和共聚焦激光扫描显微镜同时特异性差异检测定植于同一根的这些真菌的方法。该方法也可应用于其他植物-真菌相互作用的研究。用合适的荧光染料标记的特异性探针进行的植物体内 FISH 不仅可用于基础研究，还可用于检测处于潜伏阶段的定植于植物的病原真菌。
