Date Published:
2017 Jan
Publication Type:
Journal Article
Authors:
Secondary:
Mitochondrion
Volume:
32
Pagination:
16-18
PMID:
27838478
URL:
https://pubmed.ncbi.nlm.nih.gov/27838478
DOI:
10.1016/j.mito.2016.11.005
Keywords:
DNA, Mitochondrial;Hydrolysis;Mitochondrial Diseases;Polymerase Chain Reaction;Sensitivity and Specificity;Sonication;Specimen Handling;Time Factors
Abstract:
<p>Droplet digital PCR (ddPCR) is a quantitative assay that requires DNA fragmentation to maximize reaction efficiency. Here, we measured the proportion of mitochondrial DNA (mtDNA) carrying the "common deletion," a rare event, to compare quantification sensitivities between alternative DNA fragmentation methods (sonication and QIAshredder spin columns) against enzymatic digestion (traditionally used). QIAshredder showed the highest sensitivity when compared to sonication, followed by digestion. Also, both sonication and QIAshredder fragmentation had shorter processing times than enzymatic digestion; therefore, QIAshredder fragmentation and sonication are alternative DNA processing methods that maximize ddPCR quantification for the detection of rare events.</p>