X-linked agammaglobulinaemia and the underlying genetics in two kindreds

Objective: Molecular analysis of the Bruton's tyrosine kinase (Btk) gene in two unrelated families, with a combined total of seven boys, affected by X -linked agammaglobulinaemia (XLA). Methods: Protein electrophoresis and western blotting were used for the exa mination of Btk protein synthesis in blood leucocytes. Isolation of the cod ing sequence of the Btk gene was performed by amplification using the rever se transcription-polymerase chain reaction (RT-PCR) technique. Sequence alt erations were screened for by the single-stranded conformation polymorphism (SSCP) method and characterized by standard sequencing protocols. Results: Western blotting revealed Btk protein to be absent in leucocytes o f affected males from both families. A novel 3 b.p. deletion in exon 3 of t he Btk gene was found to be responsible for the XLA phenotype in the affect ed proband in one family (kindred I). A diagnostic PCR assay was establishe d to detect this mutation in other affected male siblings and carrier femal es. For the second family (kindred II), the coding sequence of the Btk gene and the promoter region were found to be normal. Conclusions: The present study has demonstrated genetic heterogeneity in th e Btk gene in South African XLA patients and has identified a novel mutatio n in this gene in the largest of the affected kindreds. The gene mutation i n the second kindred was undetermined and may be indicative of a defect in some other gene associated with Btk function or stability. Western blotting was found to be informative in establishing a deficiency of Btk protein in both probands and is recommended as a frontline procedure in the molecular diagnosis and work-up of XLA.