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. Traditionally the stability of RNA and DNA structures has been experimentally studied by UV melting curve analysis, differential scanning calorimetry and hybridization with oligonucleotide probes. Here we report a new hybridization-based approach that requires reduced amounts of target nucleic acids and inexpensive probes. In this method a series of deoxyribozyme probes produce fluorescent signal that depends on the binding affinity of a sensor to the fragment of a nucleic acid target. The sensor response enables quantitative assessment of the stability of the analyzed structure. To prove the concept, we analyzed the stability of a fragment of E.coli 23S rRNA and its DNA analog and compared the determined parameters with those theoretically predicted. The new approach allows isothermal analysis of nucleic acid structures at the concentrations as low as 0.3 nM in a cost efficient format. D. M. Kolpashchikov, Chembiochem, 2007, 8, 2039. Y. V. Gerasimova, E. Cornett, D. M. Kolpashchikov, Chembiochem, 2010, 11, 81. Y. V. Gerasimova, E. M. Cornett, E. Edwards, X. Su, K. H. Rohde, D. M. Kolpashchikov, ChemBioChem, 2013, 14, 2087-2090. Y. V. Gerasimova, D. M. Kolpashchikov. Angew. Chem. Int. Ed. Engl. 2013 52, 10586-10588. Funding from NSF CCF (1117205 and 1423219) is greatly appreciated.

General method for analysis of nucleic acid structures by deoxyribozyme sensors

251st National Meeting (2016)

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