Israelstam, Melanie Bernice Judith2014-03-192014-03-192014-03-19http://hdl.handle.net10539/14227Thesis (M.Sc.)--University of the Witwatersrand, Faculty of Health Sciences, 1998.Retinoblastoma (RB) is a malignant intraocular tum our that affects newborns and young children. The disease results from inactivation of both alleles o f the Rb-1 tumour-suppressor gene within chromosome band 1 3q 14 by deletion, mutation or gene silencing. About 4 0 % of RB cases are of the hereditary type and about 6 0 % are sporadic. The incidence in the Caucasoid population is 1 : 1 2 0 0 0 . Various mutations have been found throughout the 180 kb gene, ranging from point mutations to large deletions, but no "hot spot" or specific region for mutations has been identified. However, many mutations appear to occur as a result of C -» T transitions at CpG dinucleotides. The promoter region and exon 1 of the Rb-1 gene encompass a CpG rich region. Although CpG islands are not methylated in normal tissues, they have been demonstrated to become methylated in various tumour types. In this study, DNA from paraffin-embedded tum our specimens of southern African Negroid RB individuals was analyzed using a II methylation-specific PCR-based method, targeting specific methylation sites in order to ascertain differences of methylation patterns in RB tum ours as opposed to normal retinal tissue. Methylation of certain CpG sites in the 5 ' region of Rb-1 may result in transcriptional silencing and thereby contribute to loss of function of the gene. All of the tumours analysed were methylated at a minimum o f one site, while DNA from the normal fetal retina was unmethylated at all of the methylation sensitive CpG sites analyzed. The study provided an effective means for screening methylation changes in the Rb-1 gene that had occurred during tumorigenesis, as well as the methylation pattern of normal retinal tissue.enThesis