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Issue 18, 2011
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Real-time electrochemical monitoring of isothermal helicase-dependent amplification of nucleic acids

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Abstract

We described an electrochemical method to monitor in real-time the isothermal helicase-dependent amplification of nucleic acids. The principle of detection is simple and well-adapted to the development of portable, easy-to-use and inexpensive nucleic acids detection technologies. It consists of monitoring a decrease in the electrochemical current response of a reporter DNA intercalating redox probe during the isothermal DNA amplification. The method offers the possibility to quantitatively analyze target nucleic acids in less than one hour at a single constant temperature, and to perform at the end of the isothermal amplification a DNA melt curve analysis for differentiating between specific and non-specific amplifications. To illustrate the potentialities of this approach for the development of a simple, robust and low-cost instrument with high throughput capability, the method was validated with an electrochemical system capable of monitoring up to 48 real-time isothermal HDA reactions simultaneously in a disposable microplate consisting of 48-electrochemical microwells. Results obtained with this approach are comparable to that obtained with a well-established but more sophisticated and expensive fluorescence-based method. This makes for a promising alternative detection method not only for real-time isothermal helicase-dependent amplification of nucleic acid, but also for other isothermal DNA amplification strategies.

Graphical abstract: Real-time electrochemical monitoring of isothermal helicase-dependent amplification of nucleic acids

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Supplementary files

Article information


Submitted
08 Apr 2011
Accepted
24 Jun 2011
First published
27 Jul 2011

Analyst, 2011,136, 3635-3642
Article type
Paper

Real-time electrochemical monitoring of isothermal helicase-dependent amplification of nucleic acids

F. Kivlehan, F. Mavré, L. Talini, B. Limoges and D. Marchal, Analyst, 2011, 136, 3635
DOI: 10.1039/C1AN15289K

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