Visualization of Protein Coding, Long Non-coding and Nuclear RNAs by FISH in Sections of Shoot Apical Meristems and Developing Flowers

In addition to transcriptional regulation, gene expression is further modulated through mRNA spatiotemporal distribution, by RNA movement between cells, and RNA localization within cells. Here, we have adapted RNA fluorescence in situ hybridization (FISH) to explore RNA localization in Arabidopsis thaliana. We show that RNA FISH on sectioned material can be applied to investigate the tissue and sub-cellular localization of meristem and flower development genes, cell cycle transcripts, and plant long non-coding RNAs. We also developed double RNA FISH to dissect the co-expression of different mRNAs at the shoot apex, and nuclear-cytoplasmic separation of cell cycle gene transcripts in dividing cells. By coupling RNA FISH with fluorescence immunocytochemistry, we further demonstrate that a gene's mRNA and protein may be simultaneously detected, for example revealing uniform distribution of PIN-FORMED1 (PIN1) mRNA and polar localization of PIN1 protein in the same cells. Therefore, our method enables the visualization of gene expression at both transcriptional and translational levels with subcellular spatial resolution, opening up the possibility of systematically tracking the dynamics of RNA molecules and their cognate proteins in plant cells.