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Multiplex Detection of Gene Expression in the Intact Drosophila Brain Using Expansion-Assisted Iterative Fluorescence In Situ Hybridization
Published on: May 2, 2025
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Quantitative mRNA imaging throughout the entire Drosophila brain
Xi Long1, Jennifer Colonell2, Allan M Wong3
1Transcription Imaging Consortium, Howard Hughes Medical Institute Janelia Research Campus, Ashburn, Virginia, USA.
Nature Methods
|June 6, 2017
Summary
We developed a fast, amplification-free method for detecting single messenger RNAs (smFISH) in Drosophila brains. This technique allows precise quantification of gene expression at the subcellular level using standard or advanced microscopy.
Area of Science:
- Neuroscience
- Molecular Biology
- Genetics
Background:
- Understanding gene expression patterns is crucial for deciphering brain function.
- Current methods for mRNA detection in whole-mount tissues can be time-consuming or lack resolution.
Purpose of the Study:
- To develop and validate a rapid, high-resolution method for single-mRNA detection in intact Drosophila brains.
- To enable accurate quantification of mRNA expression with subcellular precision.
Main Methods:
- Single-molecule fluorescence in situ hybridization (smFISH) applied to cleared whole-mount adult Drosophila brains.
- Utilized standard confocal microscopy for initial validation.
- Employed a custom Bessel beam structured illumination microscope (BB-SIM) for whole-brain single-mRNA detection.
Main Results:
- Demonstrated a rapid and multiplexable smFISH approach for Drosophila brains.
- Achieved subcellular resolution for mRNA localization and abundance quantification.
- Successfully detected single mRNAs across the entire brain using BB-SIM.
Conclusions:
- The developed smFISH method offers a powerful tool for analyzing gene expression in the Drosophila brain.
- This technique facilitates accurate mRNA quantification without molecular amplification.
- The approach is compatible with both standard and advanced microscopy techniques.
