A Catalytic Hairpin Assembly-Based Approach for RNA Imaging in Living Cells.

The advancement of nucleic acid nanotechnology has resulted in broad applications of DNA- and RNA-based molecular sensors for bioanalysis. Catalytic hairpin assembly is such a type of programmable and enzyme-free nucleic acid circuit that has been popularly used in developing biosensors. Genetically encodable fluorogenic RNA-based devices have recently gained a lot of attentions as a powerful tool for intracellular imaging. Combining these two techniques together, we have developed a genetically encodable RNA-based catalytic hairpin assembly circuit for the highly sensitive detection of low-abundance RNAs in living cells. In this system, the binding of one target RNA can catalytically trigger the generation of tens-to-hundreds of fluorogenic RNA reporters and produce a significantly enhanced fluorescence signal. Here, we will introduce the detailed design, optimization, and imaging protocol of this new type of powerful imaging tools.