A dissertation submitted to the Faculty of Science, University of the
Witwatersrand, in fulfillment of the requirements for the degree of Master of
Science
Johannesburg, August 2012 / It is known that infectious agents elicit different responses in different individuals
which strengthens the view that susceptibility and resistance to infectious diseases
has a genetic component. These differences in susceptibility to disease can be
observed in populations. APOBEC3G is a member of the cytidine deaminase
gene family located on chromosome 22. It is crucial in non-permissive cells as it
functions as part of the innate immunity system and is an inhibitor of the HIV-1
accessory protein vif.
The goal of the study was to develop genotyping assays and estimate allele
frequencies. Thus, genetic variation within APOBEC3G was identified and
characterized in black South Africans. Indirect genotyping assays were designed
to amplify regions within the upstream non-coding region, and in exon 4 of the
coding region of the gene. Selected polymorphisms were then genotyped using
allele-specific PCR, RFLP-PCR and Pyrosequencing™ assays.
Reanalysis of sequence data from 2003 showed numerous SNPs were well
represented. Comparison of sequence data at various SNPs showed that allele
frequencies were similar to frequencies in other African populations. The only
sequenced SNP that deviated from the frequencies in Ensembl was -590. Thus the
sequencing was a useful tool for detection of variation. ASA proved to be the least
reliable genotyping technique as the minor allele frequency of -571 (0.59)
deviated from the published frequency of 0.894 in Africans. RFLP analysis
proved more reliable for genotyping -571 and H186R. The minor allele frequency
was estimated to be 0.84 and 0.32 for -571 and H186R respectively. The
frequency of H186R is similar to published data from An et al (2004) and Reddy
et al (2010). If SNPs are in LD they occur together on the same haplotype more
often than by chance. Usually SNPs that are in LD are in close proximity.
However our data suggests -571 and H186R SNPs which are 5kb apart are not in
LD. A LD map of chromosome 22 shows highly variable pattern of LD (Dawson
et al, 2002). Widespread regions of nearly complete LD up to 804 kb in length are
intermingled with regions of little or uundetectable LD. Haplotype analysis
showed the most frequent haplotype was GA. This was the most frequent
haplotype when the sample types were subdivided according to spoken language.
in comparison to studies from An et al, (2004) D’ of the two SNPs was estimated
at 0.967. The linkage disequilibrium (LD) revealed a non-independence of allele
segregation because the loci analyzed were strongly linked in the Apobec 3 G
gene. The data are consistent with greater genetic diversity of African populations
and can form the basis for further evaluation of the role of variation in this gene in
response to HIV.
| Identifer | oai:union.ndltd.org:netd.ac.za/oai:union.ndltd.org:wits/oai:wiredspace.wits.ac.za:10539/12695 |
| Date | 29 April 2013 |
| Creators | Ramdin, Roshilla |
| Source Sets | South African National ETD Portal |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf, application/pdf |
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