LIGASE CHAIN-REACTION ASSAY FOR HUMAN MUTATIONS - THE SICKLE-CELL BY LCR ASSAY

We can detect the beta-globin gene sickle cell mutation by using an as say based on the ligase chain reaction. The simultaneous amplification of the human growth hormone gene in the same reaction serves as a con trol for the amount of template DNA or amplification efficiency. Ligat ion products, which are biotinylated at one end and tagged with an arb itrary ''tail'' sequence at the other, are captured by hybridization t o ''tail''-complementary oligonucleotides immobilized on polystyrene m icrowells. The captured ligation products are detected colorimetricall y by use of streptavidin-alkaline phosphatase conjugate. In a study of 24 subjects, the assay unequivocally discriminated among normal, carr ier, and sickle cell genotypes.