ABSTRACT
Abnormalities in γ-globin gene regulation accounting for cases of hereditary persistence of fetal hemoglobin have been defined by polymerase chain reaction (PCR)-allele-specific oligonucleotide analysis to arise from base changes in the Aγ promoter. Sequential PCR amplification with two pairs of primers has been applied in testing unfertilized human oocytes and the first polar bodiesisolated from them. The genetic defect in the β-globin gene responsible for sickle cell disease and β-thalassemia could be diagnosed. The potential drawbacks of restriction endonuclease analysis (REA) for the SS mutation are incomplete digestion of the sample and plasmid contamination. Initially, indirect linkage analysis has been used, whereas later direct identification of mutations was developed using REA and oligonucleotide probes, and most recently, the diagnosis became possible by PCR. The original PCR amplification procedure was combined with the analysis of the β-globin amplified product by solution hybridization with specific oligonucleotide probes and subsequent digestion with a restriction enzyme to determine the genotype.
