Rapid identification of unknown heteroplasmic mutations across the entire human mitochondrial genome with mismatch-specific Surveyor Nuclease.

摘要: Mitochondrial DNA (mtDNA) mutations are responsible for mitochondrial diseases in numerous patients. But, until now, no rapid method was available the identification of unknown deleterious point mutations. Here, we describe a new strategy heteroplasmic mtDNA using mismatch-specific Surveyor Nuclease. This protocol involves following three steps: (i) PCR amplification entire human genome 17 overlapping fragments; (ii) localization mismatch(es) after digestion amplicons by Nuclease; and (iii) mutation sequencing region containing mismatch. Nuclease-based allows systematic screening to identify within 2 days. It represents an important diagnostic approach that can be routinely used molecular laboratories.