Fast mitochondrial DNA isolation from mammalian cells for next-generation sequencing

Wilber Quispe-Tintaya, Ryan R. White, Vasily N. Popov, Jan Vijg, Alexander Y. Maslov

Research output: Contribution to journalArticlepeer-review

46 Scopus citations


Standard methods for mitochondrial DNA (mtDNA) extraction do not provide the level of enrichment for mtDNA sufficient for direct sequencing and must be followed by long-range-PCR amplification, which can bias the sequencing results. Here, we describe a fast, cost-effective, and reliable method for preparation of mtDNA enriched samples from eukaryotic cells ready for direct sequencing. Our protocol utilizes a conventional miniprep kit, paramagnetic bead-based purification, and an optional, limited PCR amplification of mtDNA. Te first two steps alone provide more than 2000-fold enrichment for mtDNA when compared with total cellular DNA (~200-fold in comparison with current commercially available kits) as demonstrated by real-time PCR. Te percentage of sequencing reads aligned to mtDNA was about 22% for non-amplified samples and greater than 99% for samples subjected to 10 cycles of long-range-PCR with mtDNA specific primers.

Original languageEnglish (US)
Pages (from-to)133-136
Number of pages4
Issue number3
StatePublished - Sep 2013


  • Mitochondrial DNA
  • Next-generation sequencing

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry, Genetics and Molecular Biology(all)


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