Electronic Detection of Single-Base Mismatches in DNA with Ferrocene-Modified Probes
Abstract
Genotyping and gene-expression monitoring is critical to the study of the association between genetics and drug response (pharmacogenomics) and the association of sequence variation with heritable phenotypes. Recently, we developed an entirely electronic method for the detection of DNA hybridization events by the site-specific incorporation of ferrocenyl derivatives into DNA oligonucleotides. To perform rapid and accurate point mutation detection employing this methodology, two types of metal-containing signaling probes with varying redox potentials are required. In this report we describe a new ferrocene-containing phosphoramidite 9 that provides a range of detectable redox potentials. Using automated DNA/RNA synthesis techniques the two ferrocenyl complexes were inserted at various positions along oligonucleotide probes. Thermal stability analysis of these metal-containing DNA oligonucleotides indicates that incorporation of 9 resulted in no destabilization of the duplex. A mixture of oligonucleotides containing compounds 9 and I was analyzed by alternating current voltammetry (ACV) monitored at the 1st harmonic. The data demonstrate that the two ferrocenyl oligonucleotide derivatives can be distinguished electrochemically. A CMS-DNA array was prepared on an array of gold electrodes on a printed circuit board substrate with a self-assembled mixed monolayer, coupled to an electronic detection system. Experiments for the detection of a single-base match utilizing two signaling probes were carried out. The results demonstrate that rapid and accurate detection of a single-base mismatch can be achieved by using these dual-signaling probes on CMS-DNA chips.
Additional Information
© 2001 American Chemical Society. Received 4 January 2001. Published online 23 October 2001. Published in print 1 November 2001. We wish to thank Prof. Yitzhak Tor for helpful discussions during the preparation of this manuscript and Dr. Jost Viemetter for acquiring T_m data. We are indebted to the DNA-Chip production team at Motorola Clinical MicroSensors for preparing the chips used in this study.Additional details
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- 75737
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- CaltechAUTHORS:20170405-114813362
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2017-04-05Created from EPrint's datestamp field
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2021-11-15Created from EPrint's last_modified field