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Analysis of Mitochondrial DNA Copy Number and Its Regulation Through DNA Methylation of POLGA

  • Xin Sun
  • William Lee
  • Vijesh Vaghjiani
  • Justin C. St. JohnEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1351)

Abstract

Replication of mitochondrial DNA (mtDNA) is important for ensuring that cells have sufficient mtDNA copy number to meet their specific requirements for the generation of cellular energy through oxidative phosphorylation. A number of transcription and replication factors are required for this process, with a key factor being the nuclear-encoded mtDNA-specific DNA polymerase γ. DNA polymerase γ has a catalytic subunit (POLGA), whose gene has been shown to be DNA methylated at exon 2. This methylation is considered to be one of the key mechanisms that regulate mtDNA copy number. These findings have made it of great importance to establish optimal methods for investigating the effects of DNA methylation on mtDNA replication. Here, we provide methods to determine the extent of DNA methylation at exon 2 of POLGA as well as other gene targets of interest. We also show how mtDNA copy number is assessed and, from these two outputs, define the efficiency of mtDNA replication by calculating the mtDNA-replicative efficiency index.

Key words

mtDNA copy number MeDIP DNA methylation POLGA 

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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Xin Sun
    • 1
    • 2
  • William Lee
    • 1
    • 2
  • Vijesh Vaghjiani
    • 1
    • 2
  • Justin C. St. John
    • 1
    • 2
    Email author
  1. 1.Mitochondrial Genetics Group, Centre for Genetic DiseasesHudson Institute of Medical ResearchClaytonAustralia
  2. 2.The Department of Molecular and Translational ScienceMonash UniversityClaytonAustralia

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