Molecular prenatal diagnosis of alpha-thalassemia using real-time and multiplex polymerase chain reaction methods

Molecular analysis of two fetuses at high risk of alpha-thalassemia (alpha-thal), and their family members, was performed using real-time polymerase chain reaction (PCR) with SYBR Green 1 (SYBR-PCR) dye combined with dissociation curve analysis and multiplex PCR (m-PCR) and DNA sequencing techniques. The genotype of the fetus from one family was --SEA/--SEA (Southeast Asian deletion), which produces hydrops fetalis syndrome. The genotype of the parents was --SEA/alphaalpha. A boy with Hb H disease and his sibling fetus from the other family had the genotype --SEA/alphaCSalpha [the Hb Constant Spring (CS) mutation: alpha142, Term-->Gln (TAA>CAA in alpha2)] and alphaalpha/alphaalpha (normal), respectively. The diagnosis, based on SYBR-PCR combined with dissociation curve analysis, was in agreement with the results from the m-PCR method. This indicates that these are alternative and reliable assays for the molecular diagnosis of deletional alpha-thal.