Quantification of mitochondrial DNA copy number in suspected cancer patients by a well optimized ddPCR method

Publication date: Available online 31 August 2017 Source:Biomolecular Detection and Quantification Author(s): Ashfaque A. Memon, Bengt Zöller, Anna Hedelius, Xiao Wang, Emelie Stenman, Jan Sundquist, Kristina Sundquist Changes in mitochondrial DNA (mtDNA) content is a useful clinical biomarker for various diseases, however results are controversial as several analytical factors can affect measurement of mtDNA. MtDNA is often quantified by taking ratio between a target mitochondrial gene and a reference nuclear gene (mtDNA/nDNA) using quantitative real time PCR often on two separate experiments. It measures relative levels by using external calibrator which may not be comparable across laboratories. We have developed and optimized a droplet digital PCR (ddPCR) based method for quantification of absolute copy number of both mtDNA and nDNA gene in whole blood. Finally, the role of mtDNA in suspected cancer patients referred to a cancer diagnostic center was investigated. Analytical factors which can result in false quantification of mtDNA have been optimized and both target and reference have been quantified simultaneously with intra- and inter-assay coefficient variances as 3.1% and 4.2% respectively. Quantification of mtDNA show that compared to controls, solid tumors (but not hematologic malignancies) and other diseases had significantly lower copy number of mtDNA. Higher mtDNA (highest quartile) was associated with a significantly lower risk of both solid tumors an...
Source: Biomolecular Detection and Quantification - Category: Molecular Biology Source Type: research