General Method for Analysis of Nucleic Acid Structures by

Deoxyribozyme Sensors
Rebekah Karadeemaa and Dmitry M. Kolpashchikova,b,c
a

Chemistry Department, bBurnett School of Biomedical Sciences, cNational

Center for Forensic Science, University of Central Florida, Orlando, FL, 32816
(USA)

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.

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Funding from NSF CCF (1117205 and 1423219) is greatly appreciated.