Single transcript imaging to assay gene expression in wholemount Drosophila melanogaster tissues.

Single molecule Fluorescence in situ Hybridization (smFISH) for mRNA provides a powerful quantitative handle on expression from endogenous gene loci. While the method has been widely applied in cells in culture, applications to primary tissue samples remain fewer, and often use involved cryosectioning. Even apart from quantitative access to absolute transcript counts in specific tissue volumes, many other advantages of smFISH can be envisaged in tissue samples. Primary among these are the ability to report on subtle differences in expression among different cell types within a tissue, and the ability to correlate the expression from different target genes. Here, we present a modified method of smFISH applicable on various primary wholemount tissues from the fruit fly Drosophila melanogaster, and show the efficacy of the method in a variety of larval and adult tissue, and embryos. We also combine smFISH in tissue with immunofluorescence to demonstrate the possibility of capturing transcriptional and translational aspects of gene expression in the same tissue. Given the widespread use of Drosophila melanogaster as a model system in Developmental Biology and Genetics, such methods are likely to be of wide interest and could yield rich information about gene expression in tissues from this organism.