Design, Optimization and Validation of the ARMS PCR Protocol for the Rapid Diagnosis of Wilson's Disease Using a Panel of 14 Common Mutations for the European Population.

Background: Wilson's disease (WD) is an autosomal recessive inherited disorder of copper metabolism resulting from various mutations in the gene. Despite good knowledge and successful treatment options, WD is a severe disease that leads to disability, destructively affecting the quality of life of patients. Currently, none of the available laboratory tests can be considered universal and specific for the diagnosis of WD. Therefore, the introduction of genetic diagnostic methods that allow for the identification of the root cause at any stage over the course of the disease gave hope for an earlier solution of diagnostic issues in patients with WD.

Methods: A method for the genetic diagnosis of WD based on ARMS PCR, DreamTaq Green PCR Master Mix and modified primers has been developed. This method is able to detect 14 mutant alleles: p.His1069Gln, p.Glu1064Lys, p.Met769HisfsTer26, p.Gly710Ser, p.Ser744Pro, p.Ala1135GlnfsTer13, p.Arg778Leu, p.Arg1041Trp, p.Arg616Gln, p.Arg778Gly, p.Trp779*, p.Val834Asp, p.Gly943Ser and p.3222_3243+21del43.

Results: The primers for all mutations were highly specific with an absence of wild-type amplification. All the results were validated by direct DNA Sanger sequencing.

Conclusions: This fast and economical method provides coverage for the identified common mutations, thereby making ARMS PCR analysis using DreamTaq Green PCR Master Mix and modified primers feasible and attractive for large-scale routine use.