MALDI-TOF MS检测HPA-29-35w技术的建立及应用  

Establishment and application of MALDI-TOF MS technique for detecting HPA-29-35w

作　　者：陈洁润 周燕 李丽兰 钟周琳 蒋丽红 马婷婷 李恒聪 莫秋红 CHEN Jierun;ZHOU Yan;LI Lilan;ZHONG Zhoulin;JIANG Lihong;MA Tingting;LI Hengcong;MO Qiuhong(Nanning Blood Center/Nanning Institute of Blood Transfusion Medicine,Nanning,Guangxi 530003,China)

机构地区：[1]南宁中心血站/南宁输血医学研究所,南宁530003

出　　处：《重庆医学》2025年第1期101-107,113,共8页Chongqing Medical Journal

基　　金：广西壮族自治区卫生健康委员会自筹经费科研课题(Z20210821,Z20201430);广西自然科学基金面上项目(2023GXNSFAA026300);中国输血协会威高科研基金项目(CSBT-WG-2023-06)。

摘　　要：目的建立基质辅助激光解析电离飞行时间质谱(MALDI-TOF MS)基因分型检测技术,并应用于探索血小板抗原(HPA)-29-35w低频基因在广西地区献血人群中的多态性。方法确定7个目的基因突变位点在单核苷酸多态性数据库(dbSNP)中的rs号,利用MassARRAY平台在线引物设计工具Assay Design Suite(ADS)设计21条引物,构建7个插入HPA-29bb-35bb突变型序列的过表达载体。在广西地区献血者中随机选择29名献血者的标本和载体进行iPLEX Pro多重基因分型分析与质谱检测;同时,对上述标本进行测序,将测序结果与质谱检测结果进行对比验证,建立MALDI-TOF MS基因分型检测技术。随后采用该技术对588名广西地区献血者的标本进行HPA-29-35w基因分型检测和多态性分析。结果MALDI-TOF MS对HPA-29-35w基因分型检测结果与测序结果一致。HPA-29-35w基因分型结果显示,588名广西地区献血者的HPA-29-35bw基因型均为aa纯合子。结论成功建立MALDI-TOF MS对HPA-29-35w基因分型检测技术,并应用于广西地区献血人群的HPA-29-35w基因筛查。Objective To establish the genetic typing detection technique of matrix-assisted laser desorption ionization-time or flight mass spectrometry(MALDI-TOF MS),and to apply it to investigate the polymorphism of the human platelet antigen(HPA)-29-35w low-frequency gene in blood donor population from Guangxi area.Methods The RS number of 7 target gene mutation sites in dbSNP and 21 primers were designed by using Assay Design Suite(ADS)of MassARRAY platform online primer design tool.Seven overexpression vectors inserting into HPA-29-35bb mutant sequence were constructed.Twenty-nine blood donors were randomly selected among the blood donors team in Guangxi area.The samples and vectors conducted the iPLEX Pro multiple genotyping analysis and mass spectrometric detection.Meanwhile,the above samples were sequenced and the sequencing results conducted the comparison validation with the mass spectrometric detection results,then the MALDI-TOF MS genotyping detection technology was established.Then adopting this technology conducted the platelet antigen HPA-29-35W genotyping detection and polymorphism analysis in the samples from 588 blood donors in Guangxi area.Results The MALDI-TOF MS HPA genotyping detection results were consistent with the sequencing results.The HPA-29-35w genotyping results showed that theHPA-29-35w genotype of 588 blood donors in Guangxi area was aa homozygote.Conclusion The genotype detection method of MALDI-TOF MS for HPA-29-35w is successfully established and applied to the screening of HPA-29-35w gene among the blood donor population in Guangxi area.

关 键 词：基质辅助激光解吸电离飞行时间质谱 人血小板抗原 基因分型 基因多态性 

分 类 号：R440[医药卫生—诊断学]