All cells use RNA to regulate cellular processes, induce gene expression, and control cellular functions. Correct RNA processing and folding is essential to performing its function. For instance, one proposed cause of Alzheimer's disease the mis-folding of RNA. Cellular processes can be distorted or completely halted if RNAs are folded improperly. Analyzing correct RNA folding at cellular conditions is essential to accurately determine their behavior; however, current methods require high RNA concentrations or chemical reagents, conditions far from those found in native environments. A method for fast, inexpensive, quantitative, and accurate analysis of RNA secondary structures at low concentrations is needed for the research community. Here we propose a new, general method for quantitatively determining the stability of RNA hairpin loop and potentially other structures under near physiological conditions. By taking advantage of recent developments in synthetic biology and DNA nanotechnology, we designed a highly selective and sensitive deoxyribozyme sensor to produce a fluorescent response even at low RNA concentrations. Here we adopted the sensors for analysis of RNA secondary structures. The proposed method compares signal output of the deoxyribozymes bound to the RNA of interest and a truncated version of RNA. The folded RNA would have nucleotides competing for hybridization with the probe, while the truncated version of the RNA would not. This will allow for a quantitative determination of the stability of the RNA secondary structure. This sensitive method allows for quick probing of a specific hairpin loop, easy sample preparation, and allows for a seamless, inexpensive transition for analyzing different RNA species.

General method for analysis of RNA structures by deoxyribozyme sensors

249th National Meeting (2015)

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