This work describes a family with Duchenne Muscular Dystrophy (DMD) with a rare case of a symptomatic pregnant woman. The main aim was to perform prenatal molecular diagnosis to provide genetic counseling. The secondary aim was to suggest the molecular mechanisms causing the complex structural variant (cxSV) identified. To accomplish this, we used a multi-technique algorithm including segregation analysis, Multiplex Ligation-dependent Probe Amplification, PCR, X-chromosome inactivation studies, microarrays, whole genome sequencing and bioinformatics. We identified a duplication of exons 38-43 in the DMD gene in all affected and obligate carrier members, proving that this was the DMD-causing mutation. We also observed a skewed X-chromosome inactivation in the symptomatic woman that explained her symptomatology. In addition, we identified a cxSV (duplication of exons 38-43 and deletion of exons 45-54) in the affected boy. The molecular characterization and bioinformatic analyses of the breakpoint junctions allowed us to identify Double Strand Breaks stimulator motifs and suggested the replication-dependent Fork Stalling and Template Switching as the most probable mechanisms leading to the duplication. In addition, the de novo deletion might have been the result of a germline inter-chromosome non-allelic recombination involving the Non-Homologous End Joining mechanism. In conclusion, the diagnostic strategy used allowed us to provide accurate molecular diagnosis and genetic counseling. In addition, the familial molecular diagnosis together with the in-depth characterization of the cxSV helped to determine the chronology of the molecular events, and propose and understand the molecular mechanisms involved in the generation of this complex rearrangement.

中文翻译：

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一个家族中 DMD 基因复杂结构变异的分析

这项工作描述了一个患有杜氏肌营养不良症 (DMD) 的家庭，其中有一例罕见的有症状的孕妇。主要目的是进行产前分子诊断以提供遗传咨询。次要目的是提出导致识别的复杂结构变异 (cxSV) 的分子机制。为此，我们使用了多种技术算法，包括分离分析、多重连接依赖的探针扩增、PCR、X 染色体失活研究、微阵列、全基因组测序和生物信息学。我们在所有受影响和专性携带者成员的 DMD 基因中发现了重复的外显子 38-43，证明这是 DMD 引起的突变。我们还在有症状的女性中观察到了倾斜的 X 染色体失活，这解释了她的症状。此外，我们在受影响的男孩中发现了 cxSV（外显子 38-43 重复和外显子 45-54 缺失）。断点连接的分子表征和生物信息学分析使我们能够识别双链断裂刺激基序，并建议依赖复制的叉停顿和模板切换是导致复制的最可能机制。此外，从头缺失可能是涉及非同源末端连接机制的种系染色体间非等位基因重组的结果。总之，所使用的诊断策略使我们能够提供准确的分子诊断和遗传咨询。此外，家族分子诊断以及 cxSV 的深入表征有助于确定分子事件的年表，