Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.632683
Title: Development of biologically relevant assays for the detection of disease DNA using SERS
Author: Gracie, Kirsten
ISNI:       0000 0004 5362 6752
Awarding Body: University of Strathclyde
Current Institution: University of Strathclyde
Date of Award: 2014
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Abstract:
DNA is the fundamental material responsible for storing the genetic coding required for the development of all living organisms. Since its discovery, there has been an intense amount of research into biorecognition events and the detection of DNA sequences coding for specific diseases. The development of the polymerase chain reaction (PCR) involved the amplification of small quantities of DNA allowing for subsequent analysis. However, fluorescence-based methods such as PCR have their limitations, for example the difficulties encountered when detecting multiple targets simultaneously. Therefore, there is a need for novel techniques that overcome these limitations associated with fluorescence-based methods. This research involves the use of SERS for the multiplex detection of target DNA, investigating the possible interactions between fluorescent dyes and DNA and SERS analysis of G-quadruplex formations. A SERS-based detection assay was designed for the simultaneous detection of three bacterial meningitis pathogens; Neisseria meningitidis, Haemophilus influenzae and Streptococcus pneumoniae. By using SERS instead of fluorescence-based methods, multiplex detection was readily achieved and by using chemometric analysis it was the first report of pathogen quantification post-assay. To gain an understanding into interactions that can occur between fluorescent dyes (FAM and TAMRA), DNA and spermine, fluorescence and SERS studies were undertaken. Fluorescent studies gave an insight into the interactions that happen off the nanoparticle surface, while the SERS studies demonstrated the competitive interactions that occur between the nanoparticle surface and the two fluorescent dyes. These studies highlighted the consideration needed when selecting fluorescent dyes and target DNA sequences when designing a multiplex SERS assay. SERS was then applied to the detection of G-quadruplex formation. Previous reports used fluorescence-based methods, for example FID assays. Three ligands that selectively bind to and stabilise G-quadruplex DNA, previously used in fluorescence studies, were used and shown to have the ability to aggregate nanoparticles and act as Raman reporters. These ligands allowed for the design of the "on to off" SERS analysis of three G-quadruplex sequences.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.632683  DOI: Not available
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