Development of a high-resolution melting method for the screening of Wilson disease-related ATP7B gene mutations

Background: Wilson disease is an autosomal recessive inherited disorder of copper metabolism. The condition is characterized by excessive deposition of copper in many organs and tissues. The major physiologic aberration is excessive absorption of copper from the small intestine and impaired biliary copper excretion. The genetic defect is located at copper-transporting adenosine triphosphatase (ATPase) gene (ATP7B).

Methods: A high-resolution melting analysis (HRM) was designed to characterize the ATP7B hotspot mutations. Genomic DNA was extracted from peripheral blood samples from 14 patients and 50 normal controls. The 21 exons of ATP7B were screened by HRM analysis. Our methodology was confirmed by direct DNA sequencing.

Results: We have confirmed the 10 different hotspot mutations and 7 polymorphisms in the ATP7B gene, and also identified 1 newly-identified sequence variant (p.A476T) and 1 novel SNP (p.L776L) in 50 normal Taiwanese individuals. We estimate that the carrier frequency of WD in the Taiwanese population as probably 0.03.

Conclusions: HRM analysis is accepted as a rapid, accurate and low-cost method to screen ATP7B gene mutations.