Long-Read Sequencing Characterisation of Structural Variants in the Beta-Globin Locus and Their Functional Impact on Haemoglobin Switching in Nigerian Sickle Cell Disease Patients

This dissertation employs long-read sequencing to comprehensively characterise structural variants in the beta-globin gene cluster and their functional impact on haemoglobin switching in Nigerian sickle cell disease patients, developing an original structural variant map that explains clinically observed HbF heterogeneity not captured by standard genotyping. Short-read sequencing and SNP arrays fail to detect complex structural variants including inversions, deletions, and haplotype combinations that modify HbF expression and clinical severity, representing a major gap in SCD genomics. Oxford Nanopore Technologies long-read sequencing at 40x coverage was performed on 200 HbSS patients recruited from Lagos University Teaching Hospital and Aminu Kano Teaching Hospital, stratified by HbF percentile. Phase-resolved structural variant calling and haplotype assembly were conducted using SNIFFLES2 and phasing with WhatsHap. Functional impact of novel structural variants was predicted by integrating with ENCODE regulatory element annotations and validated by electrophoretic mobility shift assay for transcription factor binding disruption. A total of 47 structural variants in the locus control region and gamma-globin gene promoters were identified, of which 23 were novel and absent from current databases. Three novel variants predicted to disrupt BCL11A and ZBTB7A binding sites correlated with HbF levels above 20 percent in carriers. An original Nigerian Beta-Globin Structural Variant Atlas is proposed as a public reference resource. The dissertation establishes long-read sequencing as a transformative platform for complete HbF regulatory variant discovery in African SCD populations. Keywords: long-read sequencing, beta-globin locus, structural variants, haemoglobin switching, sickle cell disease.