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目的分析假肥大型肌营养不良患者 dystrophin 基因缺失断裂点的分子结构特点,探讨 dystrophin 基因缺失的发生机制。方法以 PCR 步移法定位2例 DMD 患者基因断裂位点,克隆其缺失连接片段并测序,通过 Pubmed 文献检索获取既往55例缺失连接片段序列资料,对以上57例缺失连接片段5′端和3′端断裂点两侧的序列进行重复序列、基质附着区、TTTAAA 序列以及基因缺失后修复方式的分析。结果 57例缺失连接片段中40.4%断裂点位于重复序列,其中主要是 L1元件和Alu 元件;36.3%断裂点在邻近基质附着区5 kb 之内的区域;15.0%断裂点两侧50 bp 的范围内发现有 TTTAAA 序列;基因缺失后修复的方式仅1例通过 Alu 元件同源连接,其余56例通过非同源末端连接进行修复,非同源末端连接以形成1～4 bp 的微小同源序列为主。结论重复序列、基质附着区、TTTAAA 序列以及非同源末端连接修复机制均在一定程度上参与 dystrophin 基因的断裂重组。dystrophin 基因缺失可能是由以上多种因素的综合作用所导致,染色体的物理结构可能在基因缺失中起主要作用。
Objective To analyze the molecular structure of dystrophin gene deletion in patients with Duchenne muscular dystrophy and to explore the mechanism of dystrophin gene deletion. Methods Two cases of DMD were genotyped by polymerase chain reaction (PCR). The missing and ligated fragments were cloned and sequenced. A total of 55 deletion fragments were obtained by Pubmed search. The deletion of 5 ’end and 3 Sequence analysis of the sequences flanking the end breakpoint, the matrix attachment region, the TTTAAA sequence, and the gene deletion pattern. RESULTS: The 40.4% of the 57 missing fragments were located in the repeat sequence, which were mainly L1 and Alu elements. The 36.3% breakpoint was within 5 kb of the adjacent substrate attachment area. The 50% TTTAAA sequence was found. Only 1 case was repaired by gene deletion and the other 56 cases were repaired by non-homologous end-joining, and the non-homologous ends were connected to form 1 ~ 4-bp mini-homologous sequences Mainly. Conclusions The repetitive sequences, stromal attachment regions, TTTAAA sequences and non-homologous end-joining repair mechanisms all play a part in the dysregulation of dystrophin gene. Dystrophin gene deletion may be caused by a combination of the above factors, the physical structure of the chromosome may play a major role in gene deletion.