DNA Cephalopod Nanostructure for Improving Hybridization Rates

Tatiana A. Molden, Caitlyn T. Niccum, and Dmitry M. Kolpashchikov

Department of Chemistry, University of Central Florida, 4111 Libra Drive, Physical Sciences 255, Orlando, FL 32816-2366

Fast binding of DNA and RNA by hybridization probes can expedite nucleic acid-based disease diagnosis, and recognition of specific RNA in living cells. Here we introduce a general approach for improving hybridization kinetic both in heterogeneous and homogeneous formats. Molecular beacon probe was attached in the middle of a DNA nonostructure, named ‘DNA cephalopod’, which was also equipped with several analyte capture fragments, named ‘tentacles’. The ‘DNA cephalopod’ increased the probe/analyte hybridization rates 7.5 and 27 times in solution and heterogeneous formats, respectively. The rate increase was more pronounced for folded analytes in the heterogeneous format. The approach improved limit of detection from 1 to 0.06 nM presumably due to concentrating the analyte in the microenvironment near the probe. When applied for detection of 16S E-coli RNA, the approach required additional RNA binding arms for unwinding secondary RNA structure and yielded 465 times increase in kinetics constant, which reduced detection time from ~ 300 h to 1.25 min. ‘DNA cephalopod’ approach can shorten time required for the nucleic acid-based diagnostics and therapy.