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

Abstract

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 ΔR_ct, the difference in charge transfer resistance before and after the addition of Zn²⁺, 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 ΔR_ct value. In addition, all ds-DNA films were studied by scanning electrochemical microscopy in the absence and presence of Zn²⁺, allowing us to distinguish a range of mismatched films from matched ds-DNA film.