Direct measurement of the melting temperature of supported DNA by electrochemical method.

Abstract

The development of biosensors based on DNA hybridization requires a more precise knowledge of the thermodynamics of the hybridization at a solid interface. In particular, the selectivity of hybridization can be affected by a lot of parameters such as the single-strand (ss)DNA density, the pH, the ionic strength or the temperature. The melting temperature, T(m), is in part a function of the ionic strength and of the temperature and therefore provides a useful variable in the control of the selectivity and sensitivity of a DNA chip. The electrochemical technique has been used to determine the T(m) values when the probe is tethered by a DNA self-assembled monolayer (SAM). We have built a special thin layer cell, which allows the recording of the cyclic voltammogram under controlled temperature conditions. T(m) has been determined by recording the thermogram (current versus temperature) of a redox indicator on a double-stranded hybrid (dsDNA) modified electrode and comparison with the corresponding ssDNA response. T(m) of supported DNA varies linearly with the ionic strength. The stability of the SAMs has been considered and comparison between T(m) in solution and on a solid support has been discussed.