PT  - JOURNAL ARTICLE
AU  - Lu Yang
AU  - Joshua S. Titlow
AU  - Darragh Ennis
AU  - Carlas Smith
AU  - Jessica Mitchell
AU  - Florence L. Young
AU  - Scott Waddell
AU  - David Ish-Horowicz
AU  - Ilan Davis
TI  - Single molecule fluorescence <em>in situ</em> hybridisation for quantitating post-transcriptional regulation in <em>Drosophila</em> brains
AID  - 10.1101/128785
DP  - 2017 Jan 01
TA  - bioRxiv
PG  - 128785
4099  - http://biorxiv.org/content/early/2017/04/21/128785.short
4100  - http://biorxiv.org/content/early/2017/04/21/128785.full
AB  - RNA in situ hybridization can be a powerful method to investigate post-transcriptional regulation, but analysis of intracellular mRNA distributions in thick, complex tissues like the brain poses significant challenges. Here, we describe the application of single-molecule fluorescent in situ hybridization (smFISH) to quantitate primary transcription and post-transcriptional regulation in whole-mount Drosophila larval and adult brains. Combining immunofluorescence and smFISH probes for different regions of a single gene, i.e., exons, 3’UTR, and introns, we show examples of a gene that is regulated post-transcriptionally and one that is regulated at the level of transcription. We also show that the method can be used to co-visualise a variety of different transcripts and proteins in neuronal stems cells as well as deep brain structures such as mushroom body neuropils. Finally, we introduce the use of smFISH as asensitivealternative to conventional antibody labelling to mark specific neural stem cell populations in the brain.