Molecular basis for D − J apanese: identification of novel DEL and D − alleles

Background and Objectives The occurrence of D − is approximately 0·5% in J apanese, but DEL in apparently D − individuals is relatively common compared with that in C aucasian populations. On the basis of molecular genetics, we examined D − Japanese blood donors. Methods A standard serological technique was used for R h 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 * C e 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 * DEL 8 (1), RHD with c.761 C > G (p. S er254 T er) (2), RHD with c.1252 T > A (p. T er418 L ysex26) (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 − J apanese 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.