DNA过氧化物酶探针在遗传性白内障基因诊断中的应用  

Application of Peroxidase DNAzyme Probe in the Detection of Causative Mutants for Congenital Cataract

作　　者：乔晨[1] 魏谭伟[1] 严明[1] QIAO Chen WEI Tanwei YAN Ming(Dept. of Ophthalmology, Zhongnan Hospital of Wuhan University, Wuhan 430071 , China)

机构地区：[1]武汉大学中南医院眼科,湖北武汉430071

出　　处：《武汉大学学报（医学版）》2017年第1期44-48,共5页Medical Journal of Wuhan University

基　　金：湖北省卫生与计划生育委员会西医类重点项目(编号:WJ2015MB110)

摘　　要：目的:建立应用于遗传性白内障基因突变筛查的DNA过氧化物酶检测方法。方法:针对已报道的遗传性白内障的基因突变位点晶状体蛋白CRYAB的p.R11H突变[CRYAB-32(G>A)]和缝隙连接蛋白50(Cx50)的p.S276F突变[CX50-827(C>T)],设计DNA过氧化物酶分子探针,建立DNA过氧化物酶探针和靶序列核苷酸反应体系。两个突变位点的检测均分为6组,即阳性对照组、阴性对照组、阳性实验组、阴性实验组以及2个空白对照组。主要观测各组与DNA过氧化物酶探针的显色反应。结果:DNA过氧化物酶探针与含CRYAB-32(G>A)及CX50-827(C>T)的阳性标本,在特定反应中结合均显现浅绿色,而与不含该突变的野生型DNA片段不反应不显色。结论:有过氧化物酶活性的DNA过氧化物酶作为一类新的颜色反应标签可应用于遗传性白内障基因突变的检测,能实现检测的可视化,简化检测硬件和操作步骤。Objective：To establish a visual method to screen mutations using peroxidase DNA probes in DNA samples from inherited cataract patients.Methods：According to the known causative mutations in two cataract families,we designed peroxidase DNA molecular probe and established the reaction system for peroxidase DNA probe and target sequence nucleotide.This research was approved by Zhongnan Hospital Research Ethics Committee and followed the tenets of the Declaration of Helsinki.And all patients were informed consent and signed it.Detection of the two mutations were divided into six groups,the two blank control groups and positive control group,negative control group,positive group,negative group.The main indicator was the chromogenic reaction of peroxidase of DNA probe between each group.Results：After reaction,the peroxidase DNA probe combined with p.R11 H mutation of crystalline CRYAB（CRYAB-32（G〉A））or p.S276 Fmutation of connexin 50（Cx50）（CX50-827（C〉T））appeared green color in the system.But it was colorless since the probe didn’t combine with the wild-type DNA fragment without mutations.Conclusion：The peroxidase DNA could be applied to detect the gene mutation of hereditary cataract by color reaction realized with the naked eye.It can greatly save the cost andsimplify the operation steps.

关 键 词：DNA过氧化物酶 遗传性白内障 突变检测 基因诊断 

分 类 号：R776.1[医药卫生—眼科]