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Electrochemical Detection of Single-Nucleotide Mismatches: Application of M-DNA

E. Abu-Irhayem, Y-T. Long, T. Sutherland, C-Z Li, H-B Kraatz and J.S. Lee
University of Saskatchewan, CA

DNA sensors, DNA mismatch, M-DNA, AC impedance

The detection of a single-nucleotide mismatch in a 20 base pair duplex DNA, using Electrochemical Impedance Spectroscopy, is presented. Mismatched DNA monolayers on Au electrodes were studied as normal duplex DNA (B-DNA) and after conversion to zinc metalated duplex DNA (M-DNA). Modeling of the impedance data to an equivalent circuit provides parameters that are useful in discriminating each monolayer configuration. The conversion to M-DNA caused a decrease in the resistance to charge transfer (RCT). Contrary to expectations, RCT for B-DNA was also found to decrease for duplexes containing a mismatch. Further, a method to form loosely-packed single-stranded (ss)-DNA monolayers, by duplex dehybridization, that are able to rehybridize to target strands, was developed. Rehybridization efficiencies were in the range of 40-70%. Under incomplete hybridization conditions, the RCT was the same for matched and mismatched duplexes under B-DNA conditions. However, the difference in RCT between B- and M-DNA, under incomplete hybridization, still provided a distinction. The RCT for a perfect duplex was 76(12) Ohm•cm2, whereas a mismatch in the middle of the sequence yielded a RCT value of 30(15) Ohm•cm2. The detection limit was measured and the impedance methodology reliably detected single DNA base pair mismatches at concentrations as low as 100 pM.

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