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Nucleic acid denaturation at an electrode surface

Nucleic acid denaturation at an electrode surface
Nucleic acid denaturation at an electrode surface
This thesis describes the development and improvement of an assay for the detection and discrimination of DNA sequences based on surface enhanced Raman spectroscopy (SERS) and electrochemically driven denaturation. A DNA helix can be immobilised at a nano-structured gold electrode and then denatured into its constituent components by driving the potential at the surface cathodic (electrochemical melting). This denaturation is typically monitored by observing changes in the SERS spectra of a reporter molecule attached to one of the two strands that comprises the helix. The ease of denaturation (defined as the potential required to denature half of the DNA at the electrode surface) was found to be directly related to the thermodynamic stability of the duplex. In addition to dsDNA structure, the effect of varying environmental conditions during an electrochemical melting experiment (pH and ionic strength) was explored.

A number of possible mechanisms for electrochemical melting have been ruled-out, including electrostatic repulsion from the electrode surface. It was found that peptide nucleic acid (an uncharged analogue of DNA) could be denatured electrochemically. Local pH changes were also ruled-out as a possible mechanism through experiments in which the surface pH and electrochemical melting were monitored simultaneously.

A method for detecting surface immobilised DNA without the need for the attachment of a synthetic label is described. This presents a significant milestone towards the development of a point-of care electrochemical melting assay, because no synthetic pre-treatment of the sample is required prior to analysis.
Johnson, Robert Peter
573df7a4-3f4c-493a-bc09-1d38bbff4cd0
Johnson, Robert Peter
573df7a4-3f4c-493a-bc09-1d38bbff4cd0
Bartlett, Philip N.
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Johnson, Robert Peter (2011) Nucleic acid denaturation at an electrode surface. University of Southampton, Chemistry, Doctoral Thesis, 153pp.

Record type: Thesis (Doctoral)

Abstract

This thesis describes the development and improvement of an assay for the detection and discrimination of DNA sequences based on surface enhanced Raman spectroscopy (SERS) and electrochemically driven denaturation. A DNA helix can be immobilised at a nano-structured gold electrode and then denatured into its constituent components by driving the potential at the surface cathodic (electrochemical melting). This denaturation is typically monitored by observing changes in the SERS spectra of a reporter molecule attached to one of the two strands that comprises the helix. The ease of denaturation (defined as the potential required to denature half of the DNA at the electrode surface) was found to be directly related to the thermodynamic stability of the duplex. In addition to dsDNA structure, the effect of varying environmental conditions during an electrochemical melting experiment (pH and ionic strength) was explored.

A number of possible mechanisms for electrochemical melting have been ruled-out, including electrostatic repulsion from the electrode surface. It was found that peptide nucleic acid (an uncharged analogue of DNA) could be denatured electrochemically. Local pH changes were also ruled-out as a possible mechanism through experiments in which the surface pH and electrochemical melting were monitored simultaneously.

A method for detecting surface immobilised DNA without the need for the attachment of a synthetic label is described. This presents a significant milestone towards the development of a point-of care electrochemical melting assay, because no synthetic pre-treatment of the sample is required prior to analysis.

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More information

Published date: 30 September 2011
Organisations: University of Southampton, Chemistry

Identifiers

Local EPrints ID: 206163
URI: http://eprints.soton.ac.uk/id/eprint/206163
PURE UUID: 5acd47e3-7f61-42c6-bcbd-51af44d829a3
ORCID for Philip N. Bartlett: ORCID iD orcid.org/0000-0002-7300-6900

Catalogue record

Date deposited: 16 Dec 2011 10:21
Last modified: 30 Jan 2020 01:26

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