Rapid identification of unknown heteroplasmic mitochondrial DNA mutations with mismatch-specific surveyor nuclease

Methods Mol Biol. 2009:554:301-13. doi: 10.1007/978-1-59745-521-3_19.


Identification of mitochondrial DNA (mtDNA) mutations is essential for diagnosis and genetic counseling of mitochondrial diseases. In this chapter, we describe a strategy for the rapid identification of heteroplasmic mtDNA mutations that can be used routinely in molecular genetic laboratories. This protocol involves the following three steps: (i) PCR amplification of the entire human mitochondrial genome with 17 overlapping PCR products, (ii) localization of mtDNA mismatch(es) after digestion of the 17 amplicons by Surveyor Nuclease, a member of a family of plant DNA endonucleases that cleave double-strand DNA at any mismatch site, and (iii) identification of the mutation by sequencing the region containing the mismatch.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Pair Mismatch / genetics*
  • DNA, Mitochondrial / genetics*
  • Deoxyribonucleases / metabolism*
  • Humans
  • Mitochondria / genetics*
  • Mitochondrial Diseases / diagnosis
  • Mitochondrial Diseases / genetics*
  • Mutation / genetics*
  • Polymerase Chain Reaction / methods


  • DNA, Mitochondrial
  • Deoxyribonucleases