Abstract
Pathogenic mitochondrial DNA (mtDNA) mutations are usually present in heteroplasmic forms that vary in concentration among different tissues. Manifestation of clinical phenotypes depends on the degree of mtDNA mutation heteroplasmy (mutation load) in affected tissues. It is therefore important to quantify the degree of mutation heteroplasmy in various tissues. In this chapter, we outline the design of allele refractory mutation system (ARMS)-based quantitative PCR (qPCR) analysis of common mtDNA point mutations, a cost-effective and sensitive single-step method to simultaneously detect and quantify heteroplasmic mtDNA point mutations.
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References
Wallace, D. C. (1999) Mitochondrial diseases in man and mouse, Science 283, 1482–1488.
Smeitink, J., van den Heuvel, L., and DiMauro, S. (2001) The genetics and pathology of oxidative phosphorylation, Nat Rev Genet 2, 342–352.
Enns, G. M., Bai, R. K., Beck, A. E., and Wong, L. J. (2006) Molecular-clinical correlations in a family with variable tissue mitochondrial DNA T8993G mutant load, Mol Genet Metab 88, 364–371.
Ashley, M. V., Laipis, P. J., and Hauswirth, W. W. (1989) Rapid segregation of heteroplasmic bovine mitochondria, Nucleic Acids Res 17, 7325–7331.
Bai, R. K., and Wong, L. J. (2004) Detection and quantification of heteroplasmic mutant mitochondrial DNA by real-time amplification refractory mutation system quantitative PCR analysis: a single-step approach, Clin Chem 50, 996–1001.
Moraes, C. T., Ricci, E., Bonilla, E., DiMauro, S., and Schon, E. A. (1992) The mitochondrial tRNA(Leu(UUR)) mutation in mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes (MELAS): genetic, biochemical, and morphological correlations in skeletal muscle, Am J Hum Genet 50, 934–949.
Newton, C. R., Graham, A., Heptinstall, L. E., Powell, S. J., Summers, C., Kalsheker, N., Smith, J. C., and Markham, A. F. (1989) Analysis of any point mutation in DNA. The amplification refractory mutation system (ARMS), Nucleic Acids Res 17, 2503–2516.
Wang, J., Venegas, V., Li, F., and Wong, L. J. Analysis of mitochondrial DNA point mutation heteroplasmy by ARMS quantitative PCR, Curr Protoc Hum Genet Chapter 19, Unit 19 16.
Liang, M. H., Johnson, D. R., and Wong, L. J. (1998) Preparation and validation of PCR-generated positive controls for diagnostic dot blotting, Clin Chem 44, 1578–1579.
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Venegas, V., Halberg, M.C. (2012). Quantification of mtDNA Mutation Heteroplasmy (ARMS qPCR). In: Wong, Ph.D., LJ. (eds) Mitochondrial Disorders. Methods in Molecular Biology, vol 837. Humana Press. https://doi.org/10.1007/978-1-61779-504-6_21
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DOI: https://doi.org/10.1007/978-1-61779-504-6_21
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