Whole‐mount in situ hybridization of mouse brain to precisely locate mRNAs via fluorescence tomography

Nucleic acids of intact biological tissues are rich in biological information. Whole-mount in situ hybridization is a powerful technique to mine the wealth of data contained in DNAs or RNAs, especially mRNAs. However, there are no simple, rapid approaches to precisely locate mRNAs in whole-mount tissues such as intact brains. By combining the penetration procedures of iDISCO with the signal amplification approach termed hybridization chain reaction, we herein developed a method for whole-brain in situ hybridization at cellular resolution. Based on fluorescence tomography instead of tissue clearing, this method provides a simple, rapid way to precisely locate mRNAs in the whole brain with cytoarchitectonic landmarks. As a proof of principle, we investigated the exact distribution of Cre mRNA in a Thy1-Cre mouse brain. We found high levels of Cre mRNA in most regions of the subcortical nuclei and the brain stem but comparatively low levels in major areas of the cerebral cortex. This method may have broad applications in studies of RNA function and its relations with diseases.