Primer Extension-DHPLC检测线粒体DNA 12SrRNA常见耳聋相关突变方法的建立~△

Establishment of a Common Deafness-Related Mutation Method for Detection of Mitochondrial DNA 12SrRNA by Primer Extension-DHPLC

李琦;刘亚青;孙晨;黄正华;田涛;戴朴;

1:南京医科大学附属儿童医院耳鼻咽喉-头颈外科

2:南京大学医学院附属儿童医院耳鼻咽喉-头颈外科

3:中国人民解放军总医院耳鼻咽喉-头颈外科

摘要
目的建立针对线粒体DNA 12SrRNA基因常见耳聋相关突变的Primer Extension-DHPLC(PE-DHPLC)基因诊断新方法。方法将包含线粒体DNA 12SrRNA 961insC、T1095C、C1494T、A1555G 4种不同突变的4例外周血DNA样本和4例正常对照标本经测序验证,PCR扩增靶序列,设计延伸引物,延伸后得到线粒体DNA 12SrRNA基因961insC、T1095C、C1494T、A1555G 4个常见突变的特异性延伸片段,用全变性高效液相色谱分析延伸片段混合物,分离图谱鉴定被检样本的基因型。结果 4例包含线粒体DNA 12SrRNA基因4种耳聋相关突变的DNA样本均扩增出包含4种突变的975 bp特征性条带,PE-DHPLC检测结果显示突变者有引物峰和突变峰两个峰,完全可以鉴别出4种突变,而4例正常对照标本仅显示单个引物峰。结论本实验建立了Primer Extension-DHPLC的线粒体相关耳聋突变分析新方法,可用于耳聋的基因诊断。
关键词
线粒体DNA 12SrRNA;耳聋;基因突变;变性高效液相色谱;引物延伸
基金项目(Foundation):
江苏省省级重点研发专项(BE2015608)
作者
李琦;刘亚青;孙晨;黄正华;田涛;戴朴;
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