The alpha-thalassemias are common genetic disorders that arise from reduced synthesis of the alpha globin chains of fetal (alpha2gamma2, HbF) and adult ( alpha2beta2, HbA) hemoglobins. At present, for a variety of technical, logistic and economic reasons, large-scale carrier screening and appropriate ante-natal detection programs have not been established for the populations in which this syndrome occurs. We have successfully established a simple, rapid, sensitive PCR assay for detecting common alpha gene deletion using DNA from fresh blood cell from alpha-thalassemia patients. The normal human ` globin gene includes the duplicated alpha genes (alpha2 and alpha1). All diploid cells have four alpha-chain gene (i.e., alpha alpha/alpha alpha). In alpha-thalassemia syndromes, alpha-globin synthesis is either diminished or absent due to either deletional or non-deletional abnormalities involving the alpha-globin genes. Deletion of hemoglobin gene alpha1 or alpha2 could be determined using three sets of primers (two sets for alpha1 and alpha2 gene respectively and one set for beta-actin as internal control) from alpha-thalassemia syndrome patients by allele-specific amplification and differential PCR. We have tested hemoglobin alpha gene status in 21 normal subjects and 74 alpha-thalassemia patients in the majority of whom the diagnosis was confirmed by standard Southern blotting techniques. The results clearly demonstrated that the number and specific alpha gene deletion in these patients could be differentiated by measuring the band intensity of target genes (alpha1, alpha2) and reference gene (beta-actin) after PCR amplification and gel electrophoresis. We are currently examining the feasibility of a color complementary assay with direct detection of dye-labeled PCR products in 96-well plates, which may permit a quicker and sensitive method to detect a gene deletion omitting gel electrophoresis.
