The use of in situ hybridisation to study the localisation of maternal mRNAs during Xenopus oogenesis

Summary

Introduction

A method for in situ hybridisation using Xenopus laevis oocytes is described in detail. Using this technique, we have studied the distribution of several localised maternal mRNAs during Xenopus oogenesis. Our results suggest that all of the RNAs tested, including mRNAs localised to either the animal or vegetal ends of oocytes, are evenly distributed during early oogenesis. Localisation begins during the middle stages of oogenesis, and full-grown oocytes demonstrate the most striking localisation.

One parameter that can be used to identify a gene whose product is developmentally important is the spatial pattern of its transcript. In situ hybridisation is a method commonly used for determining the spatial localisation of any relatively abundant gene transcript. Although this procedure cannot be used to detect low abundance mRNAs, and can be hindered by fixation artefacts, it can be a very useful method, particularly in conjunction with other techniques.

There are many experiments in which in situ hybridisation has been used to study how genes are expressed during development. For example, these experiments have been particularly informative with respect to the expression of homeobox genes of Drosophila (for review see Akam, 1987). In these cases this technique has been utilised in conjunction with the analysis of mutant phenotypes. In situ hybridisation has also been used in sea urchins with histone genes (Angerer et al., 1984; Venezky et al., 1981;Cox et al., 1984) transcripts of which are localised to the pronucleus until its breakdown following meiotic prophase.