A ClearSee-Based Clearing Protocol for 3D Visualization of Arabidopsis thaliana Embryos

Tissue clearing methods combined with confocal microscopy have been widely used for studying developmental biology. In plants, ClearSee is a reliable clearing method that is applicable to a wide range of tissues and is suitable for gene expression analysis using fluorescent reporters, but its application to the Arabidopsis thaliana embryo, a model system to study morphogenesis and pattern formation, has not been described in the original literature. Here we describe a ClearSee-based clearing protocol, which is suitable for obtaining 3D images of Arabidopsis thaliana embryos. The method consists of embryo dissection, fixation, washing, clearing, and cell wall staining, and enables high quality 3D imaging of embryo morphology and expression of a fluorescent reporter with the cellular resolution.