Optimised detection of mitochondrial DNA strand breaks

Intrinsic and extrinsic factors that induce cellular oxidative stress damage tissue integrity and promote ageing, resulting in accumulative strand breaks to the mitochondrial DNA (mtDNA) genome. Limited repair mechanisms and close proximity to superoxide generation make mtDNA a prominent biomarker of oxidative damage. Using human DNA we describe an optimised long-range qPCR methodology that sensitively detects mtDNA strand breaks relative to a suite of short mitochondrial and nuclear DNA housekeeping amplicons, which control for any variation in mtDNA copy number. An application is demonstrated by detecting 16-36-fold mtDNA damage in human skin cells induced by hydrogen peroxide and solar simulated radiation.