A simple and versatile cell wall staining protocol to study plant reproduction

KEY MESSAGE: The optical brightener SCRI Renaissance 2200 can be used as versatile dye to study various aspects of plant reproduction by confocal laser scanning microscopy. ABSTRACT: The study of sexual reproduction of plants has traditionally relied on light microscopy in combination with a variety of staining methods. Transgenic lines that label specific cell or tissue types with fluorescent proteins in combination with confocal laser scanning microscopy were an important development to visualize gametophyte development, the fertilization process, and to follow cell differentiation in the early embryo. Staining the cell perimeter to identify surrounding tissue is often a necessary prerequisite to put the fluorescent signal in the right context. Here, we present SCRI Renaissance 2200 (SR2200) as a versatile dye to study various aspects of plant reproduction ranging from pollen tube growth, guidance and reception to the early patterning process in the developing embryo of Arabidopsis thaliana. Furthermore, we demonstrate that SR2200 can be combined with a wide variety of fluorescent proteins. If spectral information can be recorded, even double labeling with dyes that have very similar emission spectra such as 4′,6-diamidin-2-phenylindol (DAPI) is possible. The presented staining method can be a single, easy-to-use alternative for a range of other staining protocols commonly used for microscopic analyses in plant reproductive biology. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00497-015-0267-1) contains supplementary material, which is available to authorized users.