The presence of hundreds of copies of mitochondrial DNA (mtDNA) in each human cell poses a challenge for the complete characterization of mtDNA genomes by conventional sequencing technologies1. Here we describe digital sequencing of mtDNA genomes with the use of massively parallel sequencing-by-synthesis approaches. Although the mtDNA of human cells is considered to be homogeneous, we found widespread heterogeneity (heteroplasmy) in the mtDNA of normal human cells. Moreover, the frequency of heteroplasmic variants varied considerably between different tissues in the same individual. In addition to the variants identified in normal tissues, cancer cells harboured further homoplasmic and heteroplasmic mutations that could also be detected in patient plasma. These studies provide insights into the nature and variability of mtDNA sequences and have implications for mitochondrial processes during embryogenesis, cancer biomarker development and forensic analysis. In particular, they demonstrate that individual humans are characterized by a complex mixture of related mitochondrial genotypes rather than a single genotype.
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We thank M. Whalen, J. Ptak, L. Dobbyn and N. Silliman for expert technical assistance. This work was supported by The Virginia and D. K. Ludwig Fund for Cancer Research and by National Institutes of Health grants CA57345, CA 43460, CA 62924 and CA121113.
Author Contributions Y.H., K.W.K., B.V. and N.P. designed and performed experiments, analysed data and wrote the paper. J.W. and D.C.D. performed experiments and analysed data. C.I.-D., S.D.M. and L.A.D. provided critical materials and reagents. V.E.V. analysed data and provided input to the manuscript.
Under agreements between the Johns Hopkins University, Genzyme, Exact Sciences, Beckman, Inostics, and Invitrogen, K.W.K., B.V., D.C.D., V.E.V., N.P., and L.A.D. are entitled to a share of the royalties received by the University on sales of products related to genes or technologies described in this manuscript. The University, K.W.K. and B.V. own stock in Genzyme and K.W.K., B.V., V.E.V., N.P., and L.A.D. own stock in Inostics, both of which are subject to certain restrictions under Johns Hopkins University policy. The terms of these arrangements are being managed by the Universities in accordance with their conflict of interest policies.
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He, Y., Wu, J., Dressman, D. et al. Heteroplasmic mitochondrial DNA mutations in normal and tumour cells. Nature 464, 610–614 (2010). https://doi.org/10.1038/nature08802
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