Rh E/e genotyping by allele-specific primer amplification.
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.
Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, Amsterdam.
B H
BH
Faas
S
S
Simsek
P M
PM
Bleeker
M A
MA
Overbeeke
H T
HT
Cuijpers
A E
AE
von dem Borne
C E
CE
van der Schoot
D000483:Alleles; D000818:Animals; D000911:Antibodies, Monoclonal; D001483:Base Sequence; D017931:DNA Primers; D005838:Genotype; D006801:Humans; D051379:Mice / Q000276:immunology; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense; D010641:Phenotype; D016133:Polymerase Chain Reaction / Q000379:methods*; D012204:Rh-Hr Blood-Group System / Q000235:genetics*
D000911:Antibodies, Monoclonal; D017931:DNA Primers; D016376:Oligonucleotides, Antisense; D012204:Rh-Hr Blood-Group System