PCR-GeneScan法检测遗传性聋相关基因GJB2 235 delC和mtDNA A1555G突变

PCR-GeneScan technique for detection of the GJB2 235 delC and mtDNA A1555G mutations for hereditary hearing loss

目的:检测GJB2 235delC杂合突变和mtDNA A1555G突变。方法:对120例样本进行诊断试验,其中测序GJB2 235delC杂合突变样本16例,mtDNA A1555G突变17例。用PCR方法对目标片段进行扩增,PCR产物在3100DNA sequencer(ABI)上聚丙烯酰胺胶毛细管电泳,GeneScan、GeneMarker软件数据分析。结果:120例样本均得到检测结果,检出GJB2 235delC杂合突变样本17例,mtDNA A1555G突变17例,1例正常样本误诊为235delC杂合突变而出现假阳性。结论:PCR-GeneScan技术可以同时检测2种不同基因的突变,单管多重PCR和GeneScan荧光标记法结合是同时检测多种突变一种新的思路,而且可能是一种有效的方法。

Objective：To develop a molecular genetic assay to detect the GJB2 235 delC and mtDNA A1555G mutations simultaneously based on fluorescent labeled multiplex PCR and automatic DNA fragment analyzing techniques.Method： One hundred and twenty samples were pooled in our experiment to test the feasibility of new method.The PCRs were performed and the size fragment of PCR products were analyzed on ABI 3100 Genetic Analyzer.Data analysis were taken using the software package of GeneScan and GeneMarker.Result：Seventeen samples of DNA with 235 delC and 17 samples with A1555G were tested using this protocol.A false-positive sample without GJB2 235 delC mutation was tested.Conclusion：This assay can detect both mutations in pooled DNA tests and will be a useful tool for newborn screening and carrier screening for the hereditary hearing loss in Chinese population.