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  • Title: Specific detection of deleted mitochondrial DNA by in situ hybridization using a chimera probe.
    Author: Nakamura N, Hattori N, Tanaka M, Mizuno Y.
    Journal: Biochim Biophys Acta; 1996 Sep 11; 1308(3):215-21. PubMed ID: 8809113.
    Abstract:
    We report a new method for the specific detection of the mutant mitochondrial DNA (mtDNA) that contains the 4977-bp deletion. We designed an oligonucleotide probe that was designated the 'ATP8/ND5 Chimera' probe: its 5'- and 3'-portions correspond to the ATP8 gene and the ND5 gene, respectively, and its middle portion includes the 13-bp direct repeat sequence that flanks the 4977-bp deletion. By Southern blot analysis, this chimeric probe specifically detected the deleted mtDNA, even in the presence of both normal mtDNA and other mtDNA deletions. The specificity of the probe was further confirmed by in situ hybridization of muscle fibers from patients with Kearns-Sayer syndrome who carry the deleted DNA in the heteroplasmic state. The deleted mtDNA was markedly accumulated in cytochrome-c oxidase (COX)-deficient ragged-red fibers. In tissues where multiple deleted mtDNAs were detected, such as muscle tissues from a patient with myotonic dystrophy and from an aged individual, the in situ hybridization detected a small number of muscle fibers that contained the deleted mtDNA. These results indicate that in situ hybridization using this chimera probe is a useful and specific method for detecting a small amount of deleted mtDNA.
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