多重连接依赖探针扩增在假肥大型肌营养不良症家系基因诊断中的应用

Genetic diagnosis of Duchenne/Becker muscular dystrophy by MLPA

目的评估多重连接依赖探针扩增（multiplex ligation—dependent probe amplification，MLPA）技术在假肥大型肌营养不良症（Duchenne／Becker muscular dystrophy，DMD／BMD）患者临床诊断、携带者筛查及产前诊断中的应用。方法应用MLPA法检测DMD基因79个外显子的缺失或重复突变，DNA测序以及STR毛细管电泳与连锁分析方法进行验证及辅助诊断。结果47例患儿中7例可见重复突变，31例可见缺失突变，其中7例为单个外显子缺失。23例MLPA检测阳性的患儿的母亲中有13例为携带者。产前诊断的13例胎儿中，3例为男性胎儿患者，2例为女性胎儿携带者。结论MLPA能迅速、直接、准确地检测DMD基因外显子的缺失／重复突变。联合应用MLPA、基因测序以及连锁分析可以提高DMD／BMD基因诊断的准确率。

Objective To assess the value of multiplex ligation-dependent probe amplification （MLPA） for the genetic and prenatal diagnosis of Duchenne/Beeker muscular dystrophy （DMD/BMD）. Methods Forty seven patients clinically diagnosed or suspected with DMD/BMD were recruited. Deletion or duplication of the 79 exons of the DMD gene were detected by MLPA. PCR and sequencing were used to detect single exon deletion. MLPA was also used for identifying carriers. For cases requesting prenatal diagnosis, short tandem repeat （STR） capillary electrophoresis, linkage analysis and MLPA were applied to determine fetal DMD gene. Results Among the 47 patients, deletions and duplications encompassing one or more exons were identified in 31 and 7 cases with MLPA, respectively. Seven patients had single exon deletions. However, one of which was actually a point mutation in the probe-conjugated region and was confirmed by PCR and sequencing. Of the 23 mothers with MLPA positive sons, 13 were found to carry either deletions or duplications. Prenatal diagnosis has identified 2 male affected fetuses and 3 female carrier fetuses in the 13 eases examined, which was in conformity with linkage analysis. Conclusion Our data demonstrated that MLPA is a rapid, direct and reliable method for detecting deletions or duplications of the DMD gene. It can also indicate small changes within the sequences detected by the probe. Combing MLPA with PCR, sequencing and linkage analysis could make the genetic diagnosis of DMD/BMD more accurate.