[Generation of RHD-CE(2-9)-D allele by gene conversion in cis]

Previously a few Rh-negative individuals in Caucasian and Chinese were found existing exons 1 and 10 of RHD through genomic DNA testing. The molecular mechanisms, however, remain disputed. In this study, 2 individuals carrying RHD positive, D antigen negative allele (with exon 1 and 10 of RHD) and their mRNA were investigated by using reverse transcriptase PCR (RT-PCR) and sequencing through one pair of specific primers for 5'- and 3'-non-coding region of RHD, taking a Rh-positive (Ccee) sample as control. As a result, the control sample had a normal RHD mRNA, whereas other transcripts were detected in both RHD-positive, D antigen negative individuals, which have same length and exons as the normal RHD or RHCE mRNA. Those transcripts had the same sequences with RHD in exon 1 and exon 10, while the sequences in exons 2-9 were in concordance with RHCE(e) mRNA. It indicated that a hybrid RHD allele, RHD-CE(2-9)-D, was existed in the 2 individuals, in which the exons 2-9 of RHD were substituted by homologous RHCE(e). Thus, this allele could not code normal RhD protein and therefore resulted in Rh-negative phenotype.