13例肝豆状核变性家系致病基因突变的检测和分析

Identification and Analysis of Mutations in 13 Patients with Wilson Disease

为了研究Wilson病(wilson disease,WD)基因突变的类型和发生情况,探讨WD疾病的病因、临床特点,通过采用变性高效液相色谱(denaturing high performance liquid chromatography,DHPLC)方法结合DNA测序方法对13例无亲缘关系的患者及33位亲属进行ATP7B基因所有外显子及5′端非转录区突变检测.33例DNA标本确认11种基因突变,其中包括9种错义突变(p.R778L、p.R919G、p.T1178A、p.T977M、p.K1010Q、p.C490TERM、p.A874P、p.G943S、p.G943D),1种缺失突变(2790delCAT),1种剪切位点改变(c.1708-1G>C(intron 4+1 G>C)).在13个家系中,发现4例R778L突变,2例2790delCAT,均为杂合型突变,涉及17个等位基因,本组检出率为70.8%(17/24).DHPLC-测序法是WD疾病基因突变筛查较为快捷、全面而且敏感的方法.WD基因突变存在热点区分别为3、8、12号外显子.2790delCAT、p.K1010Q、c.1708-1G>C(intron 4+1 G>C)是ATP7B基因新突变类型,p.S406A、p.V456L、p.V1140A、p.R952K是中国人比较常见的多态类型.

To determine characters and distributions of ATP7B gene mutations and discuss the pathogenic,clinical characteristics,diagnosis and treatment progress of Wilson disease（WD）,thirty three individuals（13 WD patients） from 13 no kinship WD families were subjected to polymerase chain reactions（PCR） for all exons of ATP7B gene and 5′ UTR（un-transcribed region）.PCR products were analyzed by DHPLC（dena-turing high performance liquid chromatography） and sequencing.Eleven mutations were identified by DNA sequencing,including nine missense mutations（p.R778L,p.R919G,p.T1178A,p.T977M,p.K1010Q,p.C490TERM,p.A874V,p.G943S,p.G943D）,one deletion（p.I930del,c.2790delCAT） and four polymorphisms.The mutation p.R778L occurred in 6 patients and they were all heterozygous.The methods of PCR-DHPLC and DNA sequencing are dependable tool for screening ATP7B gene mutations.Exons 3,8,12 are the muta-tion hotspots in WD gene.The mutations c.2790delCAT,p.K1010Q,c.1708-1G〉C are new mutations of ATP7B gene found in WD patients.Through genetic analysis,SNP S406A,V456L,V1140A and R952K are common polymorphic types in Chinese.