摘要
目的:建立基于MassARRAY质谱iPLEX分析技术的人血小板抗原(human platelet antigen,HPA)基因分型方法。方法:通过文献检索HPA1-29W系统的单核苷酸多态性突变或缺失位点的信息,确定35个目的基因突变位点,按MassARRAY突变位点标示方法、引物设计固定格式和引物设计软件在线设计90条引物(正、反向扩增引物和延伸引物各30条),以12个插入HPA1-29W野生型和突变型序列的合成质粒以及50例已知HPA分型的献血者样本进行扩增和质谱检测,分析灵敏度、特异度和重复性,随机抽取15例样本的一代测序和质谱分型结果进行对比验证,建立检测方法。结果:基于MassARRAY平台的HPA基因分型的质谱检测方法与测序法检测结果相符合,灵敏度为100%,特异度为100%,重复性为100%。结论:成功建立基于MassARRAY平台的HPA1-29W系统的质谱分型方法,适合中国人群HPA分型,具有临床应用前景。
Objective:This study aims to establish a genotyping method based on the iPLEX analysis of MassARRAY platform to detect human platelet antigen(HPA). Methods:The information of single nucleotide polymorphism mutation or base deletion in HPA1-29W system was searched by literature,and 29 gene mutations were identified. According to MassARRAY mutation site labeling method,primer design fixed format and primer design software,a total of 90 primers(30 forward,30 reverse amplification primers and 30 extension primers)were designed online. The sensitivity,specificity and repeatability were analyzed using 12 synthetic plasmids which were inserted into HPA1-29W wild-type or mutant sequences and 50 samples with known HPA typing. The results of 15 randomly selected samples were screened by the first generation sequencing and mass spectrometry typing to establish the detection method. Results:The results of mass spectrometry based on MassARRY platform were consistent with that of the sequencing method.The sensitivity,specificity and repeatability rate of the mass spectrum method were 100%,100% and 100%,respectively. Conclusion:The method of HPA1-29W gene mass spectrometry based on MassARRAY platform is successfully established,which is suitable for HPA typing in Chinese population,and has clinical application prospect.
作者
周春
吕红
张若洋
陈蓉
蔡莉
刘太香
石宇鹏
洪轲
刘成成
梁文飚
ZHOU Chun;Lü Hong;ZHANG Ruoyang;CHEN Rong;CAI Li;LIU Taixiang;SHI Yupeng;HONG Ke;LIU Chengcheng;LIANG Wenbiao(Jiangsu Province Blood Center,Nanjing 210042;Jiangsu Xiansheng Medical Diagnosis Co.Ltd,Nanjing 210042,China)
出处
《南京医科大学学报（自然科学版）》
CAS
CSCD
北大核心
2021年第3期361-368,共8页
Journal of Nanjing Medical University(Natural Sciences)
基金
江苏省社会发展重点项目(BE2017757)
江苏省卫生与计划生育委员会科技项目(H201514)。
