Signaling Aptamer Optimization through Selection Using RNA-Capturing Microsphere Particles.

An RNA signaling aptamer is composed of two units: a sensing aptamer that binds the input target molecule and a working aptamer that binds the output target molecule to result in a detectable signal. A conformational change of the signaling aptamer that induces an allosteric interaction with the output target molecule in response to the input target molecule depends on a junction region, which connects the two aptamer units. Efficient and effective optimization of the junction region remains a technical challenge. In this study, we demonstrate a simple strategy for optimizing the junction region through functional RNA selection using RNA-capturing microsphere particles. From approximately 0.2 million sequence variants, a signaling aptamer that enabled intracellular detection of -adenosyl methionine with a high signal-to-noise ratio, which is approximately 2-fold higher relative fluorescence increment compared to the previously reported signaling aptamer, was obtained after single round of selection. The technology demonstrated here can be used to select RNA sequences that carry out specific functions in response to particular stimuli.