Expanded application to plant reproductive tissues of a branched DNA probe-based in situ hybridization method.

PREMISE: Detecting clear tissue- and organ-specific patterns of gene expression is key to understanding the genetic mechanisms that control plant development. In situ hybridization (ISH) of mRNA is one of the most precise, yet most challenging approaches to gene expression assays. METHODS AND RESULTS: Detection of histone H4 expression in reproductive tissues of Mimulus lewisii, a model angiosperm, was optimized using the RNAscope ISH assay. The optimized protocol was used to detect histone H4 expression in reproductive tissues of two gymnosperm species, Taxodium distichum and Juniperus virginiana, without further need for species-specific optimization. Additionally, the optimized protocol was used to detect expression of CYCLOIDEA transcription factors in M. lewisii reproductive tissues without further optimization and with results similar to those previously reported. CONCLUSIONS: The RNAscope assay can quickly and sensitively generate high-quality ISH results in reproductive tissues across a breadth of plant species.