Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia

R. Saiki, S Scharf, F Faloona, K. Mullis, G. Horn, H. Erlich, N Arnheim
1985 Science  
Recent advances in recombinant DNA technology have made possible the molecular analysis and prenatal diagnosis of several human genetic diseases. Fetal DNA obtained by aminocentesis or chorionic villus sampling can be analyzed by restriction enzyme digestion, with subsequent electrophoresis, Southern transfer, and specific hybridization to cloned gene or oligonucleotide probes. With This disease results from homozygosity of the sickle-cell allele (rS) at the 3globin gene locus. The S allele
more » ... s. The S allele differs from the wild-type allele (3A) by substitution of an A in the wild-type to a T at the second position of the sixth codon of the p chain gene, resulting in the replacement of a glutamic acid by a valine in the expressed protein. For the prenatal diagnosis of sickle cell anemia, DNA ob-The authors are in the Abstract. Two new methods were used to establish a rapid and highly sensitive prenatal diagnostic test for sickle cell anemia. The first involves the primer-mediated enzymatic amplification of specific 3-globin target sequences in genomic DNA, resulting in the exponential increase (220,000 times) of target DNA copies. In the second technique, the presence of the 3A and Ps alleles is determined by restriction endonuclease digestion of an end-labeled oligonucleotide probe hybridized in solution to the amplified 3-globin sequences. The 3-globin genotype can be determined in less than 1 day on samples containing significantly less than I microgram of genomic DNA.
doi:10.1126/science.2999980 pmid:2999980 fatcat:ulo6h6r6lzdexl3iewhi7cwtce