Sensors 2008, 8(1), 193-210; https://doi.org/10.3390/s8010193
Enzyme-Linked Electrochemical Detection of PCR-Amplified Nucleotide Sequences Using Disposable Screen-Printed Sensors. Applications in Gene Expression Monitoring
1
Institute of Biophysics, v.v.i., Academy of Sciences of the Czech Republic, Kralovopolska 135, CZ-612 65 Brno, Czech Republic
2
Department of Analytical Chemistry, University of Pardubice, Nam. Cs. Legii 565, CZ-53210, Czech Republic
*
Author to whom correspondence should be addressed.
Received: 28 December 2007 / Accepted: 7 January 2008 / Published: 21 January 2008
(This article belongs to the Special Issue Utilization of Electrochemical Sensors and Biosensors in Biochemistry and Molecular Biology)
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Abstract
Electrochemical enzyme-linked techniques for sequence-specific DNA sensingare presented. These techniques are based on attachment of streptavidin-alkalinephosphatase conjugate to biotin tags tethered to DNA immobilized at the surface ofdisposable screen-printed carbon electrodes (SPCE), followed by production andelectrochemical determination of an electroactive indicator, 1-naphthol. Via hybridizationof SPCE surface-confined target DNAs with end-biotinylated probes, highly specificdiscrimination between complementary and non-complementary nucleotide sequences wasachieved. The enzyme-linked DNA hybridization assay has been successfully applied inanalysis of PCR-amplified real genomic DNA sequences, as well as in monitoring of planttissue-specific gene expression. In addition, we present an alternative approach involvingsequence-specific incorporation of biotin-labeled nucleotides into DNA by primerextension. Introduction of multiple biotin tags per probe primer resulted in considerableenhancement of the signal intensity and improvement of the specificity of detection. View Full-TextKeywords:
electrochemical detection; enzyme-linked assay; DNA hybridization; primer extension; PCR; gene expression
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Horaková-Brazdilova, P.; Fojtova, M.; Vytras, K.; Fojta, M. Enzyme-Linked Electrochemical Detection of PCR-Amplified Nucleotide Sequences Using Disposable Screen-Printed Sensors. Applications in Gene Expression Monitoring. Sensors 2008, 8, 193-210.