Molecular basis for D− Japanese: identification of novel DEL and D− alleles.

Background and Objectives The occurrence of D− is approximately 0·5% in Japanese, but DEL in apparently D− individuals is relatively common compared with that in Caucasian populations. On the basis of molecular genetics, we examined D− Japanese blood donors. Methods A standard serological technique was used for Rh D typing, and we selected 3526 D− blood samples. Genomic DNA obtained from whole blood was used for RHD analysis by polymerase chain reaction ( PCR) and sequencing. Multiplex PCR to detect all of the RHD exons and use of PCR-sequence-specific primer ( PCR- SSP) to detect RHD deletion ( RHD*01N.01) and c.1227 G> A mutation (for RHD*01 EL.01) were performed. Results Multiplex PCR and PCR- SSP revealed that 3091 of 3526 D− individuals (87·7%) were homozygous for RHD*01 N.01, and 318 individuals (9·0%) had the RHD*01 EL.01/ RHD*01 N.01 or RHD*01 EL.01/ RHD*01 EL.01 genotype. The other 103 in the 3526 individuals (2·9%) had the known D- CE- D hybrid allele, RHD*01 N.04, and the association of RHCE* Ce with RHD*01 EL.01 as well as RHD*01 N.04 was observed. The remaining 14 individuals had RHD*01 N.01 hemizygous with one of the following alleles: RHD*01 N.06 (3), RHD*01N.07 (1), RHD*04 N.01 (1), RHD* DEL8 (1), RHD with c.761 C> G (p. Ser254 Ter) (2), RHD with c.1252 T> A (p. Ter418 Lysex26) (2) and apparently common RHD (4). Adsorption and elution tests with anti- D revealed that the individuals with c.761 C> G mutation were D− while the individuals with c.1252 T> A mutation were DEL. Conclusions The RHD genotype of more than 96% of D− Japanese could be determined by conventional PCR- SSP. In addition, we identified a novel DEL allele having c.1252 T> A mutation and a novel RHD silencing allele having c.761 C> G nonsense mutation. [ABSTRACT FROM AUTHOR]

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