DIAGNOSIS OF MITOCHONDRIAL DISEASE - ASSESSMENT OF MITOCHONDRIAL-DNA HETEROPLASMY IN BLOOD

Mitochondrial DNA (mtDNA) mutations are an important cause of neurolog ical disease. The identification of causative mtDNA mutations may be p articularly troublesome in blood where there are often low levels of m utant mtDNA This is evident fi om a recent study in which heteroplasmi c mtDNA mutations in cytochrome c oxidase genes were incorrectly thoug ht to be linked to Alzheimer's disease. We wished to explore whether a nalysis of blood mtDNA, prepared by a number of DNA extraction procedu res, influenced the diagnosis of mtDNA disease. DNA was extracted by d ifferent procedures from 4 patients with heteroplasmic mtDNA mutations , and the level of heteroplasmy investigated by radioactive PCR-RFLP a nalysis. Whilst there was no consistent decrease in the level of mtDNA heteroplasmy, we observed the coamplification of a novel mtDNA pseudo gene from DNA samples extracted by a simple 'boiling' procedure using primers designed to screen for the tRNA(Leu(UUR)) A3243G mutation. Thi s pseudogene was readily amplified from DNA extracted from rho degrees (mtDNA-less) cells, confirming its nuclear location. We believe that mtDNA pseudogenes may therefore present significant difficulties in th e accurate identification of pathogenic heteroplasmic mtDNA mutations in blood, (C) 1998 Academic Press.