摘要目的：对81例假肥大型肌营养不良症(Duchenne/Becker muscular dystrophy,DMD/BMD)患者进行基因突变分析,探讨中国河南人群 DMD 基因突变的特点。方法收集81例无亲缘关系 DMD/BMD 患者的临床资料,应用多重连接依赖式探针扩增技术(multiplex ligation-dependent probe amplification,MLPA)对患者 DMD 基因进行外显子缺失/重复突变分析；对于 MLPA 检测为单个外显子缺失者,PCR 扩增相应的外显子并进行 Sanger 测序以排除假阳性；对 MLPA 检测为阴性者,应用第二代测序(next generation sequencing,NGS)技术对79个外显子及其剪切位点进行测序,并用 Sanger 测序对微小突变进行验证。结果在81例 DMD/BMD 患者中,98.8%(80/81)检测出 DMD 基因缺失、重复或微小突变。MLPA 技术检测出67例(82.7%,67/81)患者为 DMD 基因外显子缺失或重复突变,且第45~54外显子为缺失热点,其中79.1%(53/67)的患者临床表型符合“阅读框规则”；NGS 和 Sanger 测序检测出13例患者为 DMD 基因点突变,其中新突变6个,已知致病突变7个。新突变中1个为移码突变(c.4708-4709insTG),5个为无义突变(c.8812G>T、c.2131A>T、c.6035T>A、c.3426C>A、c.3055C>T)。结论本研究结果丰富了 DMD 基因突变谱；MLPA、NGS 和 Sanger 测序相结合的检测流程提高了 DMD 基因检测的敏感度和特异性,对于进一步的遗传咨询、产前诊断和基因治疗都有重要意义。

abstractsObjective To perform mutation analysis for 81 unrelated patients with Duchenne/Becker muscular dystrophy (DMD/BMD)from Henan Province.Methods Multiplex ligation-dependent probe amplification (MLPA)was used to detect potential deletion/duplications of the DMD gene.Those with single exon deletions were validated with PCR amplification and Sanger sequencing to rule out false positive results.Patients with negative MLPA results were further analyzed with next-generation sequencing (NGS),and the result was validated by Sanger sequencing.Results DMD gene deletion/duplications were detected in 67 cases by MLPA,and exons 45-54 was the most frequently deleted.The phenotypes of 79.1%patients with a deletion or duplication has conformed to the reading frame rule.In addition,13 mutations were detected by NGS and Sanger sequencing,which included 6 novel mutations including one frameshift mutation c.4708-4709insTG and 5 nonsense mutations (c.8812G>T,c.2131A>T,c.6035T>A,c.3426 C>A,and c.3055C > T).Conclusion This results have enriched the DMD gene mutation database. Combined MLPA,NGS and Sanger sequencing can greatly enhance the sensibility and specificity of genetic testing for the DMD/BMD.