Zhonghua yi xue yi chuan xue za zhi = Zhonghua yixue yichuanxue zazhi = Chinese journal of medical genetics | 2019
[Identification of ATP7B gene variant by combined use of Sanger sequencing, array CGH and quantitative PCR].
Abstract
OBJECTIVE\nTo identify the type and origin of ATP7B gene mutation in a family affected with Wilson disease by combined use of multiple methods.\n\n\nMETHODS\nPeripheral 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.\n\n\nRESULTS\nSanger 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.\n\n\nCONCLUSION\nThe 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.