摘要
目的明确1个肝豆状核变性(Wilson disease,WD)家系ATP7B基因的致病变异类型及来源,为疾病的诊断及遗传咨询提供依据.方法收集先证者及其父母和哥哥的外周血样,应用Sanger测序检测该家系4人ATP7B基因21个外显子及其侧翼序列的点突变及小片段插入/缺失,用高分辨比较基因组杂交芯片(array-based comparative genomic hybridization,aCGH)检测先证者ATP7B基因的拷贝数变异(copy number variation,CNV),并通过荧光定量PCR技术(quantitative real-time PCR,qPCR)验证该家系中另外3人ATP7B基因的拷贝数情况.结果ATP7B基因测序结果显示,先证者携带一个已知致病性杂合变异(c.2668G>A,p.V890M),该变异遗传自母亲,同时发现母亲还携带5个常见的单核苷酸多态性(single nucleotide polymorphism,SNP)位点变异,且均为杂合状态,而父亲及哥哥也携带这5个变异,所不同的是其中两个变异为纯合状态.先证者以上5个SNP均为野生型.高分辨CGH芯片结果显示,先证者ATP7B基因存在大小约4 kb的杂合缺失,包含第2、3外显子,覆盖2个SNP.qPCR结果表明,先证者父亲和哥哥ATP7B基因第2、3外显子的拷贝数约为正常对照的1/2,提示二人携带该片段的杂合性缺失;母亲结果无异常.结论本研究通过联合使用Sanger测序、CGH芯片及qPCR技术对1个WD家系进行了分子诊断,同时发现了一个新的ATP7B基因致病性CNV,丰富了ATP7B基因的突变谱.
Objective To identify the type and origin of ATP7B gene mutation in a family affected with Wilson disease by combined use of multiple methods.Methods Peripheral blood samples were collected from the proband,her parents and her brother.Sanger sequencing were used to detect point mutation and small deletion/insertion of the 21 exons and flanking sequences of the ATP7B gene in all family members.Array-based comparative genomic hybridization(aCGH)was performed to identify copy number variations(CNVs)of the ATP7B gene in the proband.The result was validated by quantitative PCR(qPCR)in other 3 members.Results Sanger sequencing indicated that the proband carried a heterozygous variation c.2668G>A(p.V890M)derived from her mother.In addition,5 common SNPs were detected in her mother,three of which were also identified in her father and brother.The 5 SNPs in the proband were of the wide type.aCGH analysis demonstrated that the proband was heterozygous for a 4 kb deletion,which encompassed exons 2 and 3 of the ATP7B gene and 2 SNPs.qPCR showed that the copy number in her father and brother was about half of the control,indicating heterozygous loss of exons 2 and 3.Conclusion The combined Sanger sequencing,array CGH and qPCR has identified a novel CNV involving the ATP7B gene.The strategy can improve the diagnostic rate for hereditary or rare diseases.
作者
徐建新
王静
王侃
许焱婷
耿娟
Xu Jianxin;Wang Jing;Wang Kan;Xu Yanting;Geng Juan(Second Department of Pediatrics,Jinhua Hospital,Zhejiang University,Jinhua,Zhejiang 321000,China;Joingenome Diagnosis,Hangzhou,Zhejiang 311188,China)
出处
《中华医学遗传学杂志》
CAS
CSCD
2019年第12期1183-1186,共4页
Chinese Journal of Medical Genetics
