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Direct detection and discrimination of double-stranded oligonucleotide corresponding to hepatitis C virus genotype 3a using an electrochemical DNA biosensor based on peptide nucleic acid and double-stranded DNA hybridization

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Abstract

Development of an electrochemical DNA biosensor for the direct detection and discrimination of double-stranded oligonucleotide (dsDNA) corresponding to hepatitis C virus genotype 3a, without its denaturation, using a gold electrode is described. The electrochemical DNA sensor relies on the modification of the gold electrode with 6-mercapto-1-hexanol and a self-assembled monolayer of 14-mer peptide nucleic acid probe, related to the hepatitis C virus genotype 3a core/E1 region. The increase of differential pulse voltammetric responses of methylene blue, upon hybridization of the self-assembled probe with the target ds-DNA to form a triplex is the principle behind the detection and discrimination. Some hybridization experiments with non-complementary oligonucleotides were carried out to assess whether the developed DNA sensor responds selectively to the ds-DNA target. Diagnostic performance of the biosensor is described and the detection limit was found to be 1.8 × 10−12 M in phosphate buffer solution, pH 7.0. The relative standard deviation of measurements of 100 pM of target ds-DNA performed with three independent probe-modified electrodes was 3.1%, indicating a remarkable reproducibility of the detection method.

Development of an electrochemical DNA biosensor for the direct detection and discrimination of double-stranded oligonucleotide (dsDNA) corresponding to hepatitis C virus genotype 3a without its denaturation using a gold electrode is described. The Electrochemical DNA sensor relies on the modification of gold electrode with 6-mercapto-1-hexanol and self-assembled monolayer of 14-mer peptide nucleic acid probe, related to the hepatitis C virus genotype 3a core/E1 region. The increase of differential pulse voltammetric responses of methylene blue, upon hybridization of the self-assembled probe with the target ds-DNA to form a triplex is the principal of the detection and discrimination. Some hybridization experiments with non-complementary oligonucleotides were carried out to assess whether the suggested DNA sensor responds selectively to the ds-DNA target. Diagnostic performance of the biosensor is described and the detection limit was found to be 1.84 × 10−12 M, in phosphate buffer solution, pH 7.0. The relative standard deviation over three independently probe modified electrodes measured at 100 pM of target ds-DNA was 3.1 % indicating a remarkable reproducibility of the detection method.

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Authors and Affiliations

  1. Electroanalytical Chemistry Laboratory, Faculty of Chemistry, University of Tabriz, Bd. 29 Bahman, 51666-14776, Tabriz, Iran

    M. H. Pournaghi-Azar & F. Ahour

  2. Faculty of Pharmacy, Tabriz University of Medical Sciences, Daneshgah St., 51664, Tabriz, Iran

    M. S. Hejazi

  3. Drug Applied Research Center & Pharmaceutical Nanotechnology Research Center, Tabriz University of Medical Sciences, Daneshgah St., 51664, Tabriz, Iran

    M. S. Hejazi

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  1. M. H. Pournaghi-Azar
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  2. F. Ahour
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  3. M. S. Hejazi
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Correspondence to M. H. Pournaghi-Azar or M. S. Hejazi.

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Pournaghi-Azar, M.H., Ahour, F. & Hejazi, M.S. Direct detection and discrimination of double-stranded oligonucleotide corresponding to hepatitis C virus genotype 3a using an electrochemical DNA biosensor based on peptide nucleic acid and double-stranded DNA hybridization. Anal Bioanal Chem 397, 3581–3587 (2010). https://doi.org/10.1007/s00216-010-3875-5

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  • DOI: https://doi.org/10.1007/s00216-010-3875-5

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