Deep sequencing unearths Nuclear mitochondrial Sequences under Leber's hereditary optic neuropathy-associated false heteroplasmic mitochondrial DNA variants

Leber’s hereditary optic neuropathy (LHON) is associated with mitochondrial DNA (mtDNA) ND mutationsthat are mostly homoplasmic. However, these mutations are not sufficient to explain the peculiar featuresof penetrance and the tissue-specific expression of the disease and are believed to be causative in associ-ation with unknown environmental or other genetic factors. Discerning between clear-cut pathogenetic var-iants, such as those that appear to be heteroplasmic, and less penetrant variants, such as the homoplasmic,remains a challenging issue that we have addressed here using next-generation sequencing approach. Weset up a protocol to quantifyMTND5 heteroplasmy levels in a family in which the proband manifests aLHON phenotype. Furthermore, to study this mtDNA haplotype, we applied the cybridization protocol. Theresults demonstrate that the mutations are mostly homoplasmic, whereas the suspected heteroplasmic fea-ture of the observed mutations is due to the co-amplification of Nuclear mitochondrial Sequences.