Identification of ATP7B gene variant by combined use of Sanger sequencing, array CGH and quantitative PCR
10.3760/cma.j.issn.1003-9406.2019.12.008
- VernacularTitle: 一个肝豆状核变性家系ATP7B基因致病性变异分析
- Author:
Jianxin XU
1
;
Jing WANG
2
;
Kan WANG
1
;
Yanting XU
2
;
Juan GENG
2
Author Information
1. Second Department of Pediatrics, Jinhua Hospital, Zhejiang University, Jinhua, Zhejiang 321000, China
2. Joingenome Diagnosis, Hangzhou, Zhejiang 311188, China
- Publication Type:Journal Article
- Keywords:
ATP7B gene;
Pathogenicity variation;
Copy number variation;
Sanger sequencing;
Array-based comparative genomic hybridization
- From:
Chinese Journal of Medical Genetics
2019;36(12):1183-1186
- CountryChina
- Language:Chinese
-
Abstract:
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.
