应用PCR-SSCP技术检测肝豆状核变性基因突变及多态

A study on mutation and polymorphism of Wilson's disease gene with polymerase chain reaction single strand conformation polymorphism

检测国人肝豆状核变性病（ＷＤ）基因外显子突变及多态。方法应用聚合酶链反应－单链构像多态性（ＰＣＲ－ＳＳＣＰ）技术分析２４个ＷＤ家系１２５名成员及５０名正常对照组外周血白细胞ＤＮＡ的ＷＤ基因第５、１４外显子（ｅｘｏｎ５，ｅｘｏｎ１４）。结果３６例ＷＤ患者有１０例可能发生ｅｘｏｎ５突变，占２７．８％；表现为４种不同的单链构像；并在正常人及ＷＤ患者均检出ｅｘｏｎ５多态。在ｅｘｏｎ１４未发现突变。结论ｅｘｏｎ５可能为国人ＷＤ基因突变热点之一。本研究对ＷＤ基因诊断及ＷＤ基因突变与多态性分析有重要应用价值。

Objective To detect the mutation and polymorphism of exons of Chinese Wilson's disease gene (WND). Method Exon 5, 14 of WND were analysed by using polymerase chain reaction single strand conformation polymorphism (PCR SSCP) technique in DNA separated from peripheral blood of 125 members of 24 WD families and 50 normal controls. Results The PCR products of exon5 and exon14 were 232bp and 301bp, respectively. There were six different mobility shift detected in exon 5 but none in exon 14. Four of these such as the “R” type detected in 4 cases; the “S” type in 1 case; the “T” type in 2 cases and the “X” type in 3 cases which might be caused by different mutation were identified in 10 of 36 WD patients (27.89%). Another two patterns might be derived from DNA polymorphism. Conclusion Exon5 might be a hot point of mutation in WD gene in Chinese. It was valuable for gene diagnosis of WD as well as analysis of the mutation and polymorphism on WD gene.