A false-positive diagnosis for the common MELAS (A3243G) mutation caused by a novel variant (A3426G) in the ND1 gene of mitochondria DNA

Background: Several mutations in mitochondrial DNA (mtDNA) are associated w ith the syndrome of mitochondrial myopathy, encephalopathy, lactic acidosis , and stroke-like episodes (MELAS). The "common" MELAS mutation, A3243G in the tRNA leucine (UUR) gene, affects approximately 80% of cases and is asso ciated with respiratory chain complex I deficiency. Methods and Results: The A3243G mutation creates an ApaI restriction endonu clease site and can be detected by polymerase chain reaction (PCR) amplific ation of a region of mtDNA containing nt 3243, followed by ApaI digestion a nd electrophoretic analysis of the resulting fragments. Analysis of mtDNA f rom a child with complex I deficiency indicated the presence of the mutatio n homoplasmically in heart, liver, and skeletal muscle. Sequencing revealed only normal tRNA leucine (UUR) sequence, and a novel variant at nt 3426 in the ND1 subunit of complex I, which creates an ApaI site. ApaI digestion r esults in fragments of similar size to those found in patients with the A32 43G mutation. Conclusions: A novel variant at nt 3426 of mtDNA creates an ApaI site and c an potentially cause a false-positive result for the presence of the A3243G mutation. Given the highly polymorphic nature of mtDNA, care must be exerc ised in choosing primers for restriction endonuclease-based diagnostic test s for point mutations, and confirmation of a mutation by an independent met hod is recommended.