Abstract
A hybridization assay procedure was devised that makes possible quantitation of the ratio of mRNA of alpha to mRNA of beta globin chains in an RNA sample. The assay uses the radioactive synthetic DNA copies obtained by incubation of RNA-dependent DNA polymerase of avian myeloblastosis virus with rabbit globin mRNA that is 80-90% enriched in mRNA specific for synthesis of alpha or beta globin chains. The rabbit alpha-chain mRNA is obtained from the postribosomal supernatant of rabbit reticulocyte lysates; the rabbit beta-chain mRNA is obtained from the largest polysomes of rabbit reticulocytes treated with L-O-methylthreonine. Sufficient homology exists between rabbit and human globin chains and globin mRNAs that the synthetic DNA copies of chain-specific rabbit globin mRNA hybridize with human globin mRNA. Applied to the study of globin mRNA isolated from reticulocytes of humans with alpha and beta thalassemia, the technique revealed marked quantitative deficiency of alpha-chain mRNA relative to beta-chain mRNA in alpha thalassemia and similar deficiency of beta-chain mRNA relative to alpha-chain mRNA in beta thalassemia. The thalassemia syndromes are therefore characterized by true quantitative deficiency of the mRNA specific for the affected globin chain.
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