title: Rapid identification of unknown heteroplasmic mitochondrial DNA mutations with mismatch-specific surveyor nuclease.
abstract: 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.
authors.affiliation: Department of Medical Genetics, Archet 2 Hospital, CHU Nice, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Bannwarth
authors.forename: Vincent
authors.initials: V
authors.lastname: Procaccio
authors.forename: Véronique
authors.initials: V
authors.lastname: Paquis-Flucklinger
mesh_terms: D020137:Base Pair Mismatch / Q000235:genetics*; D004272:DNA, Mitochondrial / Q000235:genetics*; D003851:Deoxyribonucleases / Q000378:metabolism*; D006801:Humans; D008928:Mitochondria / Q000235:genetics*; D028361:Mitochondrial Diseases / Q000175:diagnosis / Q000235:genetics*; D009154:Mutation / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods
chemical_list: D004272:DNA, Mitochondrial; D003851:Deoxyribonucleases