A rapid and sensitive PCR screening method for point mutations associated with mitochondrial encephalomyopathies

Abstract Alterations of the mitochondrial DNA, encoding important parts of the cellular energy-generating system (oxidative phosphorylation, OXPHOS), are often associated with the occurence of degenerative neuromuscular diseases. Especially point mutations in the mitochondrial tRNA genes, which cannot be complemented by the nuclear encoded tRNAs, are candidates for severe defects of the OXPHOS system. An A to G transition at nt 8344 in the tRNA Lys gene has been associated with MERRF disease whereas an A to G substitution at nt 3243 in the tRNA Leu gene has been linked to the MELAS syndrome. These two mtDNA alterations as well as point mutations in protein-coding genes can be detected simultaneously by an allele-specific amplification of the altered mtDNA. This assay allows the reliable detection of heteroplasmic point-mutations, even if the mutated DNA appears to a small extent of less than 1%.