Live FISH: imaging mRNA in living neurons.

摘要

This paper describes a novel technique to identify and study neurons that express specific mRNAs. The method is called Live FISH because it is like fluorescent in situ hybridization (FISH) but can be applied to living tissues. In Live FISH, molecular beacons, which fluoresce only in the presence of the complementary mRNA, are delivered by gene gun into living neurons. One of its many benefits over existing gene expression assays is that the neurons are alive. Because identified neurons are living, they can be targeted for subsequent single-cell assays, such as electrophysiology. In addition, dye delivered with the molecular beacons illuminates dendritic morphology. Live FISH is cheaper than generating transgenic animals, and the endogenous mRNA is assayed without covalent modification or genetic manipulation. This report demonstrates the feasibility of Live FISH with yellow fluorescent protein (YFP) mRNA in living mouse retinas, although molecular beacons can be rationally designed to any mRNA of interest.