Application of maternal plasma DNA analysis for noninvasive prenatal diagnosis of Hb E-β-thalassemia

To establish simple noninvasive prenatal diagnosis of common β-thalassemia in Southeast Asia, we have evaluated the possibility of identifying the 3 most common β-thalassemia genes [βE, β17A-T, and β41/42(-CTCC)] by analysis of fetal DNA in maternal plasma using combined conventional polymerase chain reaction (PCR) and real-time quantitative PCR. Maternal plasma was obtained from peripheral blood of Thai pregnant women collected during the first and second trimesters of gestation. DNA was prepared from 200 μL plasma using a QIAmp Blood Mini Kit. Identifications of the βE, β41/42(-CTTT), and β17A-T in plasma DNA were carried out using semi-nested (for βE) and nested (for β41/42 and β17) real-time allele-specific PCR methodologies, and the results were compared with those obtained on fetal tissue analysis with routine invasive procedure. Twenty-six fetal βE genes were correctly identified by maternal plasma DNA analysis of 39 pregnant women investigated. The fetal β41/42 and β17 mutations were detectable in 6 of 12 and 4 of 9 maternal plasma specimens, respectively, which were in concordance with the results obtained by routine invasive procedure. The noninvasive prenatal diagnostic methods developed should potentially prove useful for detection of paternally inherited mutation and for providing the exclusion of pregnancies at risk for this common genetic disorder in the region.