Detection of the M268T Angiotensinogen A3B2 mutation gene based on screen-printed electrodes modified with a nanocomposite: application to human genomic samples

Abstract

We report on a stable and uniform nanocomposite (NC) as an enhancing element on the surface of a screen printed electrode for the detection of the M268T mutation of Angiotensinogen gene by DNA hybridization and DNA synthesis. This NC consists of multiwalled carbon nanotubes, pyrenebutyric acid (PBA) and chitosan. Two kinds of DNA biosensors were constructed by covalently coupling amino modified oligonucleotides to the carboxylic groups of PBA. Methylene Blue (MB) was employed as an electroactive probe for the detection of DNA. The two DNA biosensors were applied to detect the complementary sequence by differential pulse voltammetry. The results suggested that the peak currents of MB on the two biosensors are linearly related to the logarithm of the concentrations of target DNA in the 1.0 aM to 10 nM and in the 1 aM to 0.1 nM ranges, with detection limits of 0.11 and 0.24 aM for normal and mutant DNA, respectively. The selectivity experiment also showed the biosensors to be able to distinguish between target DNA and non-complementary sequences, and between normal homozygote, mutant homozygote and heterozygote. The biosensor was applied to quantify the products of PCR amplification of the Angiotensinogen gene (that is related to Atherosclerosis) extracted from human blood samples and gave satisfactory results. We expect this scheme to possess potential application in the detection of other genes.