Detection of achondroplasia G380R mutation from PCR amplicons by using inosine modified carbon electrodes based on electrochemical DNA chip technology

Abstract

The detection of Achondroplasia G380R mutation from real samples was performed by monitoring the oxidation signal of guanine. Inosine-substituted 12-mer capture probes related to the homozygous or heterozygous alleles of Achondroplasia G380R mutation were adsorbed onto carbon paste electrode (CPE) surface. No guanine signal was obtained from the capture probes, since the inosine bases were electroinactive. Then, these probes were hybridized with the denatured PCR samples on the CPE surface. The hybridization between the probe and target sequences was determined by using the oxidation signal of guanine in connection with differential pulse voltammetry (DPV). The oxidation signal of guanine was observed as a result of the specific hybridization between the probe and PCR sample. The changes in the peak height of the guanine signal provided the information whether the PCR sample contained heterozygous allele or homozygous allele. Numerous factors affecting the hybridization and nonspecific binding events were optimized to detect down to 41.24 fmol/ml target DNA. The electrochemical detection of Achondroplasia G380R mutation from PCR samples greatly shortened and simplified the diagnosis for the homozygous mutant type, which is lethal for the newborn. (C) 2003 Elsevier B.V. All rights reserved.