Screening for clinically significant non-deletional alpha thalassaemia mutations by pyrosequencing

Anna Haywood (1) , Helene Dreau (1) , Adele Timbs (1) , Anna Schuh (1) , John Old (1) , Shirley Henderson (1)

1 National Haemoglobinopathy Reference Laboratory, Molecular Haematology, Haemophilia Centre, Churchill Hospital

Non-deletional α-thalassaemia is associated with a higher degree of morbidity and mortality than deletional forms of α-thalassaemia. Screening for the common deletional forms of α-thalassaemia by Gap-PCR is widely practiced; however, the detection of non-deletional α-thalassaemia mutations is technically more labour-intensive and expensive, as it requires DNA sequencing. In addition, the presence of four very closely homologous alpha globin genes and the frequent co-existence of deletional forms of α-thalassaemia present another layer of complexity in the detection of these mutations. With growing evidence that non-deletional α-thalassaemia is relatively common in the UK, there is a demand for technologies which can quickly and accurately screen for these mutations. We describe a method utilising pyrosequencing for detecting the ten most common clinically significant non-deletional α-thalassaemia mutations in the UK. We tested 105 patients with non-deletional α-thalassaemia and found 100% concordance with known genotype as identified by Sanger sequencing. We found pyrosequencing to be simpler, more robust, quicker, and cheaper than conventional sequencing, making it a good choice for rapid and cost-effective diagnosis of patients with suspected non-deletional α-thalassaemia. The technique is also likely to help expedite prenatal diagnosis of pregnancies at risk of α-thalassaemia major.

Mots clés: Alpha thalassaemia; Pyrosequencing; Haemoglobinopathy screening; Prenatal diagnosis

Anna Haywood, Helene Dreau, Adele Timbs, Anna Schuh, John Old, et al.. Screening for clinically significant non-deletional alpha thalassaemia mutations by pyrosequencing. Annals of Hematology, 2010, 89 (12), pp.1215-1221. ⟨10.1007/s00277-010-1013-2⟩. ⟨hal-00549426⟩