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目的 研究我国X连锁显性遗传性肾炎一家系 (又名Alport综合征 ,Alport’ssyndrome ,AS)编码Ⅳ型胶原α5链的COL4A5基因突变特点。方法 提取 3 5例现存家系成员基因组DNA ,根据COL4A5基因中与外显子相邻的内含子序列设计引物 ,分别作DNA聚合酶链式反应 (PCR) ,将该基因所有 5 1个外显子扩增。以琼脂糖凝胶电泳鉴定产物 ,作单链构象多态性 (SSCP)分析检测PCR产物 ,对迁移率异常者进行DNA直接测序。结果 PCR -SSCP分析发现 17个外显子PCR产物单链DNA迁移率异常 ,通过DNA直接测序发现 9个外显子可疑突变 ,但反向测序均未能证实。结论 该家系COL4A5基因外显子未发生突变 ,其突变可能发生在该基因内含子或其它致病基因。
Objective To study the mutation of COL4A5 gene encoding type Ⅳ collagen α5 chain in a Chinese family with X-linked dominant genetic nephritis (Alport syndrome, Alport’s syndrome, AS). Methods The genomic DNA of 35 members of the existing family members was extracted. According to the sequence of introns in exon of COL4A5 gene, primers were designed and used for PCR respectively. All 5 1 exons Sub-amplification. The products were identified by agarose gel electrophoresis, and single-strand conformation polymorphism (SSCP) analysis was used to detect the PCR products. DNA sequencing was performed on those with abnormal mobility. Results The results of PCR-SSCP analysis showed that there were 17 exon PCR products with abnormal single-stranded DNA. The direct sequencing of DNA revealed 9 suspicious mutations in exon, but none of them was confirmed by reverse sequencing. Conclusion There is no mutation in the exon of COL4A5 gene in this pedigree. The mutation may occur in the gene introns or other pathogenic genes.