Study On The RHD Gene Polymorphs In Chinese

The Rh blood group system is one of the most polymorphic blood group systems in humans. The clinically importance is inferior to ABO blood group system. D antigen has the strongest antigenicity among all the Rh antigens. In particular, the highly immunogenic D antigen is involved in haemolytic disease of the newborn (HDN) and transfusion reactions. D antigen is encoded by RHD gene. Because the gene has polymorphs at molecule level, it will influence the D antigen density.In the study, blood samples were collected from 441 D-negative donors at the Shanghai blood center between July 2004 and October 2005. 74 D-negative blood samples from Xinjiang in September 2004 were also collected. All samples were determined the Rh phenotypes by using monoclonal anti C, c, E, e reagents in saline. Then the blood samples were tested by using the indirect antiglobulin test (IAT) to exclude the Weak D and Partial D phenotype. The rest samples were retested by anti-D adsorption and elution assay to screen the DEL. Then the genomic DNA of DELs was isolated by using QIAamp DNA blood mini kit.Multiplex PCR was used to determine whether the DEL has the intact RHD gene. Then sequence-specific primer polymerase chain reaction (PCR-SSP) was used to explore the nucleotide mutations of RHD alleles 885T and 1227A. 7 DEL samples were additional analysed by genomic DNA sequencing. 2 samples were further analysed by cDNA sequencing.Using adsorption and elution technique, 72 DELs were found among 441 Shanghai D-negative samples, 7 DELs, include 1 Chinese Uigur, 1 Chinese Hui and 5 Chinese Han, were found among 74 Xinjiang D-negative samples. DEL phenotype is highly linked to C antigen. In my study, only 1 DEL didn't carry C antigen.Detected by MPX PCR and PCR-SSP, all 79 DELs had intact RHD gene. 75 DELs, including 1 Chinese Uigur and 1 Chinese Hui samples, carried RHD 1227A allele. None of the samples had the RHD 885T allele. 4 DELs might have novel RHD alleles.7 DEL samples, including 4 specimens may have novel RHD alleles, and 1 Chinese Uigur, 1 Chinese Hui, 1 Chinese Han samples detected the 1227A allele by PCR-SSP, were sequenced to study the RHD genomic DNA of exonl~10. The sequencing results confirmed that PCR-SSP can accurately detected the 1227A allele. New RHD alleles were found among other 4 samples. 1 DEL had a RHD gene 251 T>C mutation, 1 DEL had a RHD gene 3 G>A mutation, 1 DEL had a RHD gene 53...