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Rh E/e genotyping by allele-specific primer amplification

1995; Elsevier BV; Volume: 85; Issue: 3 Linguagem: Inglês

10.1182/blood.v85.3.829.bloodjournal853829

1528-0020

B. H. W. Faas, Suat Şimşek, PM Bleeker, M.A.M. Overbeeke, H. Theo M. Cuijpers, AE von dem Borne, CE van der Schoot,

Blood disorders and treatments

It has been shown that the Rhesus (Rh) blood group antigens are encoded by two homologous genes: the Rh D gene and the Rh CcEe gene. The Rh CcEe gene encodes different peptides: the Rh C, c, E, and e polypeptides. Only one nucleotide difference has been found between the alleles encoding the Rh E and the Rh e antigen polypeptides. It is a C-- >G transition at nucleotide position 676, which leads to an amino acid substitution from proline to alanine in the Rh e-carrying polypeptide. Here we present an allele-specific primer amplification (ASPA) method to determine the Rh E and Rh e genotypes. In one polymerase chain reaction, the sense primer had a 3′-end nucleotide specific for the cytosine at position 676 of the Rh E allele. In another reaction, a sense primer was used with a 3′-end nucleotide specific for the guanine at position 676 of the Rh e allele and the Rh D gene, whereas the antisense primer had a 3′-end nucleotide specific for the adenine at position 787 of the Rh CcEe gene. We tested DNA samples from 158 normal donors (including non-Caucasian donors and donors with rare Rh phenotypes) in these assays. There was full concordance with the results of serologic Rh E/e phenotyping. Thus, we may conclude that the ASPA approach leads to a simple and reliable method to determine the Rh E/e genotype. This can be useful in Rh E/e genotyping of fetuses and/or in cases in which no red blood cells are available for serotyping. Moreover, our results confirm the proposed association between the cytosine/guanine polymorphism at position 676 and the Rh E/e phenotype.