https://www.um.edu.mt/library/oar/handle/123456789/34117 2026-08-12T07:35:16Z https://www.um.edu.mt/library/oar/handle/123456789/34702 Title: The effects of trans-regulatory loci on globin gene control Abstract: Augmenting the level of foetal haemoglobin (HbF) in sickle cell disease or β thalassemia patients would greatly ameliorate the symptoms associated with these diseases. This can only be achieved by first understanding the genetic switch from foetal to adult haemoglobin that is commonly referred to as γ to β globin gene switching. In this study a combination of clinical research coupled with basic research was carried out. One hundred seven β-thalassemia heterozygotes were identified by the screening program carried out by the Laboratory of Molecular Genetics, University of Malta. A complete blood count accompanied all blood samples to the laboratory. HbF and HbA2 measurements were conducted using a High- Pressure Liquid Chromatography. DNA sequencing of all samples was performed for the β globin gene and its promoter. The results showed the commonly encountered ~ thalassaemia alleles in the Maltese population; these being IVSI-6C, IVSI-llOA, Codon 39 T, IVSII-1A and other rare ones as well including a deletion of 2 nts in the β globin gene coding sequence identified for the first time in Malta. The -158 C-7T 5'Gγ globin SNP genotyping was performed by XmnI restriction enzyme digest. Three polymorphisms (rs766432, rs1l886868 and rs4671393) in BCLllA and another three (rs4895441, rs28384513 and rs9399137) in MYB were genotyped using Real Time PCR. There was a positive association between the MYB and HbF levels in β° Codon 39 heterozygotes whilst not with other β thalassaemia alleles. Only one dimorphism in BCLI1A was associated with higher HbF levels in β thalassaemia heterozygotes. As exemplified by EKLF in previous work carried out in the laboratory, and other transcription factors such as BCLIIA and MYB in this study are thought to play a significant role in promoter-specific gene activation and warrant further investigation regarding their role in globin switching. Description: M.SC.BIOMED.SCI. 2012-01-01T00:00:00Z https://www.um.edu.mt/library/oar/handle/123456789/34268 Title: The further development of DNA pooling for gene discovery and public health genomics Abstract: A model based on quantitative haemoglobin (Hb) genetics was explored to seek alleles that may have been "trans-selected" by beta thalassaemia. Hb is a duplex of two heterodimers (or 2αβ). Quantitative data on Rb variants such as Rb S, in the context of different genotypes, suggested a model that could be suitable to account for the broad range of severity in various complex diseases that are caused by multiple al1eles with quantitative effects. Interplay between two to three alleles at two to three loci (α,β,γ) could be associated with levels of a heterodimeric molecule such as Hb S, from as little as 5% to as much as 100% of total Hb. A search for other heterodimers was conducted based on the assumption that like Hb, duplexes of heterodimers, having subunits coded on non-syntenic chromosomes could display expression profiles over a broad range of values due to interplay of subunits at the level of protein assembly. Five heterodimers involved in inflammation, having one of their sub units coded by either chromosome 11 or 16 were selected from databases. Fifteen single nucleotide polymorphisms (SNPs) were quantified in the random (neonate) Maltese population and in adult β thalassaemia heterozygotes. Two of the single nucleotide variants (Integrin alpha-M (ITGAM) c.3436C>T and ITGAM c.2573C>T), which were in linkage disequilibrium (LD) with each other, resulted in odds ratios of 1.3 in the comparison between beta thalassaemia heterozygotes and controls. The two SNPs are located on chromosome 16 in the gene coding for the ITGAM subunit of the ITGAM/beta-2 heterodimer. The beta 2 (β2) gene is located on chromosome 21. Furthermore, whole exome sequencing on the Maltese random neonate DNA pool (n=50) gave a strong linear correlation (R² = 0.9066) between known Maltese allele frequencies and allele frequencies obtained from whole exome sequencing. Sequencing of DNA pools is a well-suited technique for quantification of allele frequencies and discovery of other alleles that may have been subjected to trans-selection. ITGAM/β2 is coded for by non-syntenic genes. Therefore selection pressure on one of the sub units may have been transmitted to the other as in the case of the alpha and beta globin subunits of Hb. Description: M.SC.PATHOLOGY 2012-01-01T00:00:00Z