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Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods

Includes bibliographical references.

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Main Author: Loubser, A S
Format: Thesis
Language:English
Published: Division of Virology 2015
Subjects:
Virology
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access_status_str Open Access
author Loubser, A S
author_browse Loubser, A S
author_facet Loubser, A S
author_sort Loubser, A S
collection Thesis
description Includes bibliographical references.
format Thesis
id oai:open.uct.ac.za:11427/13400
institution University of Cape Town (South Africa)
language eng
last_indexed 2026-06-10T12:34:33.896Z
license_str Not specified — see source repository
provenance_str_mv Harvested via OAI-PMH from UCTD — University of Cape Town Open Access Repository
publishDate 2015
publishDateRange 2015
publishDateSort 2015
publisher Division of Virology
publisherStr Division of Virology
record_format dspace
source_str UCTD — University of Cape Town Open Access Repository
spelling oai:open.uct.ac.za:11427/13400 Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods Loubser, A S Virology Includes bibliographical references. Immune system pressure on HIV-1 replication drives the antigenic changes seen over time. The monitoring of changes in viral sequences can provide important information on the nature of the immune response and the correlates of protection. Viral diversification may also occur due to other selective pressures such as cell availability and differences in viral fitness. Information on the genetic characteristics of HIV-1 variants present in the mother and her infected infant are useful data for establishing whether any common features exist between source infection and transmitted genotypes. This helps in the understanding of the mechanism of transmission and the selective pressures occurring during and following transmission. The overall aim of this study was to explore alternative methods other than DNA sequencing for the monitoring of genetic diversity in the third variable region (V3) of the HIV-1 env gene of integrated HIV-I variants in peripheral blood mononuclear cells (PBMC's) derived from infected mother-child pairs. Two methods for displaying DNA differences were compared: I-leteroduplex Mobility Assay (I-IMA) and Base Excision Sequence Scanning (BESS). These methods were validated using sequence data. Extracted PBMC DNA from infected mother-child pairs were used to amplify the V3 region by nested PCR. DNA fragments were cloned into plasmid vectors and analyzed by HMA and BESS to establish subtype and intrasample genetic diversity. In addition, a PCR-ELISA quantitation system was developed to measure copy numbers of integrated HIV-1 genomes in order to confirm whether a sufficient number of template molecules were present to be representative of the total viral quasispecies. In conclusion, this study compared two methods (HMA and BESS) as cost-effective alternatives to DNA sequencing for HIV-1 diversity studies. in addition, a novel application of the BESS assay was demonstrated. Diversity studies are reliant on estimation of adequate input of amplifiable copies. The PCR-ELISA quantitation system developed provided an efficient and specific method for determining DNA copy number. 2015-07-14T08:34:13Z 2015-07-14T08:34:13Z 2004 Master Thesis Masters MSc http://hdl.handle.net/11427/13400 eng application/pdf Division of Virology Faculty of Health Sciences University of Cape Town
spellingShingle Virology
Loubser, A S
Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods
thesis_degree_str Master's
title Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods
title_full Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods
title_fullStr Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods
title_full_unstemmed Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods
title_short Genetic diversity of subtype C HIV-1 env variants in peripheral blood mononuclear cell (PBMC) DNA from infected mother-child pairs : a comparison of Heteroduplex Mobility Assay (HMA) and Base Excision Sequence Scanning (BESS) methods
title_sort genetic diversity of subtype c hiv 1 env variants in peripheral blood mononuclear cell pbmc dna from infected mother child pairs a comparison of heteroduplex mobility assay hma and base excision sequence scanning bess methods
topic Virology
url http://hdl.handle.net/11427/13400
work_keys_str_mv AT loubseras geneticdiversityofsubtypechiv1envvariantsinperipheralbloodmononuclearcellpbmcdnafrominfectedmotherchildpairsacomparisonofheteroduplexmobilityassayhmaandbaseexcisionsequencescanningbessmethods

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