Browsing Masters Degrees (Genetics) by browse.metadata.advisor "Burger, Nicolaas Francois Visser"
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- ItemComparing the methylomes of two genealogically linked Russian wheat aphid biotypes using whole genome bisulfite sequencing(Stellenbosch : Stellenbosch University, 2020-03) du Preez, Pieter Herodus; Botha-Oberholster, Anna-Maria; Burger, Nicolaas Francois Visser; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.ENGLISH ABSTRACT: Diuraphis noxia (Kurdjumov, Hemiptera: Aphididae), commonly known as the Russian wheat aphid (RWA), is an economically important cereal pest. Although it does not spread any plant viruses, the severe symptoms caused by RWA feeding poses a significant threat to world wheat production. Commercial wheat cultivars, resistant to RWA, have been developed and are effective at preventing yield losses. However, new, more virulent RWA biotypes (morphologically similar aphid populations, with the ability to successfully feed on previously resistant cultivars) are continuously emerging, leading to a breakdown in resistance. The molecular mechanisms driving biotypification (the development of new biotypes) have not been identified yet. It has been proposed that an epigenetic modification, such as DNA methylation, might a possible means whereby biotypification might occur. The aim of this study was to explore the possible link between levels of DNA methylation and virulence in the RWA, by performing a whole genome bisulfite sequencing (WGBS) analyses on the South African biotypes SA1 and SAM. Together, SA1 and SAM form a good model for the study of virulence as they are closely related, yet at opposite ends of the virulence scale. The overall trends in RWA DNA methylation, observed in this study, correlates with what has previously been reported in insects: genic bodies, especially exons, are the most methylated regions in the genome, with most of the methylation occurring at CpG sites. The ratio of observed to expected CpG sites in a region has been used to infer levels of methylation, as increased methylation has been correlated to a decrease in CpG abundancy; In this study, however, no correlation was found between CpG abundancy and DNA methylation level. This technique, therefore, is not applicable for insect genes. Using the generated WGBS data, 148 genes were found to be differentially methylated between the two biotypes. The relative expression of five of these genes, which were selected based on gene ontology and the degree of differential methylation, along with that of DNA methyltransferase 3 (DNMT3) and ten-eleven translocation enzyme (TET), was quantified and compared between biotypes at 0, 6, and 48-hours after performing host-shifts from susceptible to resistant host plants. DNMT3 is the enzyme responsible for the establishment of DNA methylation, while the TET enzyme catalyses the first step of the process of demethylation. The time points were selected to correlate with defence responses elicited from the host plants. While no clear pattern could be observed in the differences in relative expression of differentially methylated genes between biotypes, or within a biotype between time points, a major increase in expression of both DNMT3 and TET 6-hours after performing a host shift to the resistant cultivar Tugela Dn5 was observed in SAM. This seems to indicate an increased rate of methylation, demethylation, or both methylation and demethylation in SAM, while under stress, which might contribute to its increased ability to respond to, and overcome, plant defence responses.