Development of a real-time PCR incorporating high resolution melting analysis to screen HIV-1 samples for resistance-related codons

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dc.contributor.authorSacks, David
dc.date.accessioned2011-02-01T09:47:16Z
dc.date.available2011-02-01T09:47:16Z
dc.date.issued2011-02-01
dc.descriptionMSc (Med), Virology, Faculty of Health Sciences, University of the Witwatersranden_US
dc.description.abstractIntroduction High resolution melting analysis (HRMA) accurately, rapidly and cost effectively detects single nucleotide polymorphisms by monitoring DNA dissociation kinetics. This technology was applied to HIV samples to assess whether it could be used to detect clinically relevant drug resistance mutations. Methods HRMA-PCR assays incorporating unlabeled probes were designed to genotype 12 mutation codons in the HIV-1 p66/p51 of engineered plasmids and 116 HIV-1 samples. Results HRMA correctly genotyped 63%-88% of the K103N, Y181C, M184V, Q151M and G190A mutations. Each assay had a 1.7%-3.4% discordance, most of which was due to the increased analytical sensitivity of HRMA (~5-20%). Only mutant K65R and V106M were correctly identified while the 41, 67, 70, 215 and 225 codons could not be genotyped. Assay modifications had some success in masking the affects of polymorphisms. Conclusion These assays can be used for genotyping selected major HIV-1 resistance mutations and should be further developed as a resistance surveillance tool.en_US
dc.identifier.urihttp://hdl.handle.net/10539/8993
dc.language.isoenen_US
dc.subjectHIV-1en_US
dc.subjectmutationsen_US
dc.subjectdrug resistanceen_US
dc.titleDevelopment of a real-time PCR incorporating high resolution melting analysis to screen HIV-1 samples for resistance-related codonsen_US
dc.typeThesisen_US
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