Rapid differentiation of five common alpha-thalassemia genotypes by polymerase chain reaction.
The alpha-thalassemias are common genetic disorders that arise from reduced synthesis of the alpha-globin chains. At present, large-scale carrier screening and clinically valuable antenatal detection programs have not been established for the congenital disorder alpha-thalassemia (alpha-thal). We have developed a simple nonradioactive polymerase chain reaction (PCR) approach that can detect and differentiate several common alpha-globin gene deletional alpha-thals regardless of the break points. When three primer sets were used--two gene-specific sets for the alpha1- and alpha2-globin genes and one set for the beta-actin gene (serving as an internal control)--PCR products from genomic DNA were simultaneously amplified and analyzed after coamplification and gel electrophoresis. The number of alpha-globin genes present in the subjects was determined by the intensity of alpha1 and alpha2 bands normalized with that of beta-actin when using densitometry. Our results demonstrate that five common genotypes of deletional alpha-thal are differentiated by the ratios of alpha1/beta-actin and alpha2/beta-actin. We also examined the feasibility of coupling this allele-specific amplification to a color-complementary assay. This easy and reproducible PCR assay is suitable for identifying alpha-thal carriers in screenings of large populations and improving genetic counseling.
Tang
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DC
Molecular and Clinical Hematology Branch, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA
Fucharoen
S
S
Ding
I
I
Rodgers
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GP
D000199:Actins / Q000235:genetics; D001483:Base Sequence; D017931:DNA Primers; D005838:Genotype; D005914:Globins / Q000235:genetics; D006801:Humans; D016133:Polymerase Chain Reaction; D017085:alpha-Thalassemia / Q000235:genetics*
D000199:Actins; D017931:DNA Primers; D005914:Globins