Advantages and limitations of in situ hybridisation as exemplified by the molecular genetic analysis of Drosophila development

Summary

Introduction

Over the past five years the application of in situ hybridisation to the analysis of gene expression has made an immense contribution to the study of Drosophila development (for reviews, see Akam, 1987; Ingham, 1988). The advent of this technique not only allowed the verification of the inferences about normal gene expression based upon classical genetic analysis, but has also facilitated the investigation of regulatory interactions between genes. In addition, in a number of cases, in situ analysis has revealed novel roles for genes not previously predicted by mutational analysis.

In the first part of this paper we outline briefly the methodology and applications of the various in situ hybridisation protocols which have been used with this organism. In the second part, we review the recent advances in the analysis of Drosophila development and discuss the uses and limitations of the in situ technique in the study of genes of unknown biochemical function.

Methodology

Several protocols have been developed for the analysis of transcripts in situ (Hafen et at, 1983; Akam, 1983; Ingham, Howard & Ish-Horowicz, 1985; Mahoney & Lengyel, 1987), but probably the most convenient and widely applicable of these employs tissues which have been embedded and sectioned in paraffin wax.

A major advantage of wax-embedded material over frozen tissue is the ease with which it can be handled and sectioned; large amounts of material can be accumulated and stored at different stages of the procedure, either prior to or following embedding. In addition, wax sections can be cut on most microtomes, obviating the need for expensive (and cumbersome) cryostats, an important consideration where money and space are limiting.