多重PCR-CE用于血小板抗原HPA-2、-3、-15基因分型的研究

Genotyping of human platelet antigens HPA-2,-3 and -15 by multiplex PCR-CE

关键词:  血小板抗原  基因分型  多重PCR  毛细管电泳

Key words:  human platelet antigens  genotyping  multiplex PCR  capillary electrophoresis

摘要:

［目的］建立一个用于血小板抗原HPA -2、-3、-15基因分型的多重PCR和毛细管电泳（CE）方法，用其调查大连地区汉族人群HPA -2、-3、-15系统的基因频率。［方法］分别设计HPA -2、-3、-15等位基因序列特异性引物和一条公共引物，用荧光标记公共引物。多重PCR法扩增HPA -2、-3、-15。然后用毛细管电泳检测扩增产物，并根据产物片段长度的差异进行基因分型。将多重PCR-CE与常规PCR-SSP方法相比较。将获得的大连地区汉族人群HPA -2、-3、-15系统基因分型数据与国内其它地区比较。［结果］多重PCR-CE和PCR-SSP的基因分型结果完全一致。大连地区汉族人群HPA-2a、-2b、-3a、-3b、-15a、-15b基因频率分别为0.9200、0.0800、0.5750、0.4250、0.5800和0.4200；与国内山东、上海、浙江汉族人群相比，HPA -2、-3、-15等位基因频率差异无显著性意义（P>0.05）；与新疆维吾尔族人群相比，HPA-2、-3等位基因频率差异无显著性意义（P>0.05），但HPA-15等位基因频率差异有显著性意义（P<0.05）。［结论］HPA多重PCR-CE基因分型方法快速、简单、准确；国内HPA-15等位基因分布存在着种族差异。

Abstract:

［Objective］To develop a multiplex PCR-CE method for genotyping of human platelet antigens(HPA)-2,-3 and -15 and investigate gene frequencies of HPA-2,-3 and -15 in Dalian Han population with the method.［Methods］The allele-specific primers and common primers for genotyping of HPA-2,-3 and -15 were designed.The common primers were labeled with fluorescence.The genes of HPA-2,-3 and -15 were amplified using multiplex PCR.Then PCR products were detected using CE.HPAs were genotyped according to different length of PCR products.The multiplex PCR-CE was compared with the conventional PCR-SSP.In addition,dates of HPAs genotyping in Dalian Han population were compared with those in other Chinese populations.［Results］There was complete concordance of results for all samples tested by multiplex PCR-CE and PCR-SSP.The gene frequencies of HPA-2a,-2b,-3a,-3b,-15a and -15b in Dalian Han population were 0.9200,0.0800,0.5750,0.4250,0.5800 and 0.4200 respectively,which did not significantly differ from those in Shan dong,Shanghai and Zhejiang Han populations (P>0.05).The gene frequencies of HPA-2,-3 in Dalian Han population did not significantly differ from those in Xinjiang Uighurs population (P>0.05),but the gene frequency of HPA-15 in Dalian Han population significantly differed from that in Xinjiang Uighurs population (P<0.05).［Conclusions］Multiplex PCR-CE for genotyping of HPAs is a fast,simple and accurate method.There is different in frequency distribution of HPA-15 alleles among different races of China.