PRENATAL-DIAGNOSIS OF THE THALASSEMIA SYNDROMES BY RAPID DNA ANALYTICAL METHODS

Prenatal diagnostic strategies applied today are based mainly on polym erase chain reaction (PCR) analytical protocols. In Greece a wide rang e of mutations underlie the thalassaemic haemoglobinopathies, and cons equently a variety of PCR-based methods are required to facilitate dia gnosis of all potential abnormal genotypes. PCR protocols include thos e which are relatively simple and others that are technically challeng ing, but very few have been designed for high through-put clinical dia gnostics. Over a period of 18 months we carried out prenatal diagnosis in 147 pregnancies (150 fetal samples) at risk for a wide range of ha emoglobinopathies. This involved the precise characterization of paren tal genotypes and the subsequent analysis of fetal DNA samples. In thi s series, 18 different mutations in the a-or P-globin clusters were id entified. For the characterization of these mutations, five PCR-based protocols were selected: denaturing gradient gel electrophoresis (DGGE ), amplification refractory mutation system (ARMS) PCR, restriction en donuclease analysis of PCR fragments, oligonucleotide hybridization an d 'gap' PCR for detection of deletions. To avoid spurious diagnosis du e to contamination of fetal samples, two additional methods were used to genotype polymorphic variable nucleotide tandem repeat (VNTR) regio ns of the genome in parental and fetal samples. Through analysis of th e results we assess the advantages and drawbacks of the selected PCR-b ased protocols for providing routine clinical diagnostics.