Molecular and family analyses revealed two novel RHD alleles in a survey of a Chinese RhD-negative population.
OBJECTIVE
RHD alleles are considered variable in the Chinese RhD-negative persons. The purpose of the present work was to elucidate the molecular bases of two novel RHD alleles identified in a survey of a Chinese RhD-negative population.


METHODS
A total of 163 RhD-negative blood samples were investigated. The sequences of RHD exons were examined by RHD exon specific multiplex polymerase chain reaction (PCR) and PCR with sequence-specific primers (PCR-SSP). Characterizations of RHD intron 2 and Rhesus box were performed by PCR-PstI digestion. The DNA and cDNA sequences of the novel alleles were determined by PCR and reverse transcriptase-PCR (RT-PCR) sequencing analysis. A family study was performed to investigate the segregation of a novel RHD allele.


RESULTS
One hundred and twenty-nine samples (79.1%) had no RHD gene. Twenty-seven samples (16.6%) carried RHD-CE-D hybrid alleles. The remainder seven samples (4.3%) appeared to have an intact RHD gene. Three of them were sequenced for RHD gene and two novel alleles, RHD 325del and RHD intron 2 1A, were identified. The deletion of a nucleotide A at position 325 in the allele RHD 325del resulted in a stop codon at amino acid position 118. The RHD intron 2 1A allele was generated from a splice mutation and its transcript sequence had no exon 2. Family study indicated that the RHD 325del allele was inherited with a Ce haplotype.


CONCLUSIONS
This study provides the molecular bases of RHD alleles RHD 325del and RHD intron 2 1A. The existences of RHD 711del, RH (D1 CE2-9 D10), and RH (D1 CE2-9 D10) alleles in the Chinese population were confirmed. A PCR-SSP-based assay for rapid detection of RHD 325del and RHD intron 2 1A alleles was established and it could be used to predict the RHD genotype in the Chinese RhD-negative subjects.
Ye
L-Y
LY
Blood Group Reference Laboratory, Shanghai Blood Center, Shanghai, China.
Guo
Z-H
ZH
Li
Q
Q
Zhu
Z-Y
ZY
D000483:Alleles; D044466:Asian People / Q000235:genetics*; D001483:Base Sequence; D005838:Genotype; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics*; D010641:Phenotype; D012204:Rh-Hr Blood-Group System / Q000235:genetics*
D012204:Rh-Hr Blood-Group System; C026410:Rho(D) antigen