REVERSE DOT-BLOT DETECTION OF THAI BETA-THALASSEMIA MUTATIONS

Pending curative therapy, newborn screening and prenatal diagnosis are essential to the management of beta thalassaemia, Diagnosis using ele ctrophoretic methods is difficult in the presence of composite phenoty pes and high Hb F levels, Direct DNA detection of mutant alleles circu mvents both problems, but the enormous diversity of beta-thalassaemia mutations poses challenges for this approach, Among PCR-based tests, t he reverse dot-blot method enables screening several mutations with a single hybridization reaction. Unfortunately it has often been targete d to only the common mutations of a particular ethnic population, nece ssitating the use of more arduous detection methods for the less commo n mutations. We developed a reverse dot-blot strip for the 10 beta-tha lassaemia mutations, including the beta-thalassaemic haemoglobinopathi es Hb E and Hb Malay, that account for 96% of beta thalassaemia in Tha iland, and another strip for six less common Thai mutations. The secon d strip precludes the need for more technically challenging methods. T o avoid problems associated with secondary structure of amplified full -length target DNA, we amplified and labelled beta-globin DNA as two s horter fragments that encompassed all known Thai mutations, Reverse do t-blotting is a rapid, accurate method for detecting beta-thalassaemia mutations.