RNA-Fluorescence In Situ Hybridization for the Detection of RNA in Mammalian Oocytes and Embryos

RNA metabolism plays an essential role in the development of both oocytes and embryos. Their dependence on stored maternal transcripts is due to a long period of silenced transcription in which the oocyte undergoes meiotic maturation and fertilization. Although maternal RNAs are unusually stable, they should be replaced by "zygotic" transcripts during the transition from oocyte to zygote. Analysis of ncRNA and mRNA distribution in the oocyte can provide clues to the fate of RNA in the single-cell environment.Our work focuses on the visualization of the subcellular distribution of specific RNAs in mammalian oocytes and early embryos. The localization of many RNAs in the oocyte and embryo is still not fully understood. In this chapter, we describe an optimized protocol for RNAscope, a type of RNA fluorescence in situ hybridization (RNA FISH), which is a valuable technique for exploring mammalian oocytes and embryos. The method is based on a specific probe design strategy that enables signal amplification together with simultaneous background suppression. Additionally, it is suitable for quantitative spatio-temporal analysis of specific RNA transcripts. RNAscope, a simple and a reliable protocol, contributes to advancing our understanding of RNA biology in the context of germ cell development.