Large DNA inversions, deletions, and TaqI site mutations in severe haemophilia A

Large DNA inversions caused by an intrachromosomal recombination between homologous regions located in intron 22 and 5' of the factor VIII gene have recently been identified in patients with severe haemophilia A. To evaluate better the prevalence of this large inversion and to estimate the overall sensitivity of the Southern blot/hybridization method we analysed the factor VIII gene of 49 unrelated patients with severe haemophilia A. All patients were screened for the inversion mutation, TaqI site mutations, and deletions. Mutations were identified in 31 (63%) patients, and comprised 24 large inversions, 4 partial deletions, and 3 point mutations. Three different haplotypes were characterised in the patients presenting the inversion mutation, confirming its independent origin. Two novel deletions are reported: a large one spanning from intron 14 to intron 22 and a deletion of 86 bp comprising the 3' region of exon 1 and 39-41 bp of intron 1. DNA sequencing of the deletion junction showed no significant homology between normal 5' and 3' sequences around the breakpoints. A novel missense mutation is also reported: CGA-->GGA, Arg-2209 to Gly. These results confirm that the inversion mutation is the most common cause of severe haemophilia A and indicate that the Southern blot/hybridization assay should be used as the first method for screening of mutations in severe haemophilia A.