| At present,30human blood group systems have been officially named by the International Society of Blood Transfusion (ISBT), comprehending nearly300blood group antigens in which the ABO and Rh blood groups as well as some rare blood groups existing in very low frequency are included. In clinical transfusion practices, despite the same ABO and Rh blood group, it is possible that the alloimmune transfusion reaction can be caused by rare blood groups incompatible between the recipients and the donors. However, existing serological test methods, limited by the deficiency or titer of the antibodies can only detect a small number of rare blood group antigens. With the development of molecular biology and its application in blood group research, it has been clarified by the molecular mechanisms of almost all blood group systems. Based on the single nucleotide polymorphism(SNP)sites of the blood group antigen alleles, sequence specific primers can be designed and then used for the genotyping of the blood group antigen alleles. This approach has been increasingly used in the screening for rare blood group attributing to its advantage of both high throughput and make-up for the shortage of monoclonal antibodies in the serological test.Multiplex PCR, a process of screening rare blood group antigens, is a technic that carried out in one reaction system added the sequence-specific primers for different blood group antigens alleles, and the interpretation of the results are based on the PCR results. However, it’s not broadly applied due to the specificity of the blood group antigens which is difficult to get the high frequency or low frequency blood group antigens samples, making it is impossible to get the appropriate alleles controls for the blood group antigens, and limits the application of blood group genotyping.Thus a multiplex PCR system, as an implement for the screening of the SNPs of the Oka, Fya and s blood group alleles, was established, and then438random donors samples were tested in which2Fy(a-) samples were found without s-and OK(a-) samples detected. Meanwhile, to validate the sensitivity and specificity of our multi PCR system, the standard plasmids, as a compared screening system to the high frequency of multiple antigen-negative controls, comprising the s and Oka blood group allele mutant SNP site, were successfully constructed on the basis of the PCR-based site-directed mutagenesis technology.With a simple polymorphism, OK blood group system is the24th blood group system defined by the International Society of Blood Transfusion (ISBT) in1999. Its only antigen is the antigen Oka, a high frequency antigen located in the immunoglobulin superfamily molecule CD147. There are rare reports on the OK (a-) phenotype both at home and abroad, and at present only eight null phenotype (OK (a-) were found in Japanese families and a Caucasian, yet other populations have not been found yet.The antigen Oka is encoded by the BSG gene with the full-length of10.8kb. It is found that the polymorphic site of the BSG gene is in the2exon. The original amino acid Glu92is replaced by Lys92in Oka antigen, as long as a single nucleotide G/A mutation occurs at the274site of the BSG gene cDNA, thus the rare OK (a-) phenotype. Due to the high frequency of the Oka antigen, there is no OK (a-) phenotype reported in China. In this paper, the method of full-length cDNA sequencing was adopted to analyze the polymorphism of BSG gene encoding the OK blood group system, and to further understand the distribution of OK blood group system in Chinese population. In total,42cases of normal random blood donors were sequenced for the full length cDNA of the BSG gene, and three polymorphisms were detected. They are cDNA195site C/T mutation,234site C/G mutation and327site T/C mutation. The result of its coding sequence analysis indicated that the three mutations does not cause the change of the amino acid sequence, and had almost no effect on its antigenicity.There are fewer studies on the role of CD147present on red blood cells so far, most of which are carried out in vivo. CD147, a highly glycosylated protein, whose glycosylation is tissue-specific, that is, in different tissues it has different degree of glycosylation due to its different functions. CD147is a single transmembrane glycoprotein present on the erythrocyte, and our Western Blotting result demonstrated that it is the high-glycosylated type with a molecular weight of55KD. Some researchers presumed that CD147has a similar function with CD47on the membrane of red cells as a signal to identify and phagocytize by macrophages, and the binding of CD147and its ligand can result in the inhibiting signal of phagocytosis. The continuous reduction of CD147on old erythrocyte membrane weakened the inhibiting signal, and then phagocytized by macrophages. To investigate this hypothesis, we detected the expression of CD147both on the young and old erythrocyte obtained by the method of capillary centrifugation using the flow cytometry. The result indicated that CD147expressed on old erythrocyte showed a significant trend of decrease, having an average decline ratio33.56%. However, further studies about the role of CD147on erythrocyte and its ligands are needed. |
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