Probing nucleobase mismatch variations by electrochemical techniques: exploring the effects of position and nature of the single-nucleotide mismatch.

Electrochemical impedance spectroscopy (EIS) has been used as an ultrasensitive tool for label-free detection of single-nucleotide mismatches in double-stranded DNA (ds-DNA) films. In this study, we have explored the effects of the position and of the type of single-nucleotide mismatch in ds-DNA on gold surfaces and were able to distinguish mismatch positions and mismatch pairs. The single-nucleotide mismatches A-C, A-A and A-G were introduced at three positions within the sequence in bottom, middle and top positions of ds-DNA, the films were studied by EIS, and the impedance results were interpreted with the help of equivalent circuits. The DeltaR(ct), the difference in charge transfer resistance before and after the addition of Zn(2+), was used to distinguish single-nucleotide mismatch within the DNA sequences. Importantly, the mismatch pair is easily distinguishable at the middle position. A purine-pyrimidine mismatch can be distinguished from purine-purine mismatch by its lower DeltaR(ct) value. In addition, all ds-DNA films were studied by scanning electrochemical microscopy in the absence and presence of Zn(2+), allowing us to distinguish a range of mismatched films from matched ds-DNA film.