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Permanent link (DOI): https://doi.org/10.7939/R3H93R

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High throughput genotyping of single nucleotide polymorphisms in the Plasmodium falciparum dhfr and dhps genes by asymmetric PCR and melt-curve analysis Open Access

Descriptions

Other title
Subject/Keyword
melt-curve analysis
Plasmodium falciparum
sulfadoxine-pyrimethamine
genotyping
DHFR
Type of item
Thesis
Degree grantor
University of Alberta
Author or creator
Cruz, Rochelle
Supervisor and department
Yanow, Stephanie (Public Health Sciences)
Kipp, Walter (Public Health Sciences)
Examining committee member and department
Yasui, Yutaka (Public Health Sciences)
Pilarski, Linda (Oncology)
Department
Department of Public Health Sciences
Specialization

Date accepted
2011-08-31T21:27:50Z
Graduation date
2011-11
Degree
Master of Science
Degree level
Master's
Abstract
Mutations within Plasmodium falciparum dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes contribute to resistance to antimalarials such as sulfadoxine-pyrimethamine (SP). Single nucleotide polymorphisms (SNPs) within codons 51, 59, 108 and 164 in the pfdhfr gene and codons 436, 437, 540, 581, and 613 in the pfdhps gene are associated with SP treatment failure. This study focused on the development of a genotyping assay based on asymmetric real-time PCR and melt-curve analysis. Probes specific to each SNP hybridize differentially to mutant and wild-type sequences, generating distinct melting curves. The analytical sensitivity and specificity were determined on the LightCycler® platform using reference strains of malaria. The assay was validated with clinical samples from patients with malaria and compared to a gold standard test. This assay was also tested for its application to in-gel PCR technology for point-of-care testing in resource-limited areas. Genotyping is possible with further enhancements to this technology.
Language
English
DOI
doi:10.7939/R3H93R
Rights
License granted by Rochelle Cruz (remnace@ualberta.ca) on 2011-08-31T19:23:49Z (GMT): Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of the above terms. The author reserves all other publication and other rights in association with the copyright in the thesis, and except as herein provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.
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