In situ localization of citrus exocortis viroid RNA using an optimized RNAscope™ assay

Due to their small size, locating pathogenic RNAs, such as viroids, in plant tissue and cell organelles has been difficult. Viroids are circular single-stranded RNA virus-like pathogens that reduce growth, vigor, yield economically important crops potato, tomato, hops citrus. Viroid infections plants largely diagnosed by dot blot hybridization, PAGE-gels, or RT-PCR. Because traditional situ hybridization studies for viroids often limited target signal amplification specificity of binding, we examined the use RNAscope™ (Advanced Cell Diagnostics Inc., Newark, CA), an method amplifies a single target, study cellular distribution citrus exocortis viroid (CEVd). We found after optimization, CEVd was localized nuclei infected cells clearly distinguishable punctates visible with light microscopy at low magnification, suggesting can be used situ.