Droplet digital PCR shows the D-Loop to be an error prone locus for mitochondrial DNA copy number determination.

ABSTRACT: Absolute quantification of mitochondrial DNA copy number (mCN) provides important insights in many fields of research including cancer, cardiovascular and reproductive health. Droplet digital PCR (ddPCR) natively reports absolute copy number, and we have developed a single-dye, multiplex assay to measure rat mCN that is accurate, precise and affordable. We demonstrate simple methods to optimize this assay and to determine nuclear reference pseudogene copy number to extend the range of mCN that can be measured with this assay. We evaluated two commonly used mitochondrial DNA reference loci to determine mCN, the ND1 gene and the D-Loop. Harnessing the absolute measures of ddPCR, we found that the D-Loop amplifies with a copy number of ~1.0-1.5 relative to other sites on the mitochondrial gen

SUBMITTER: Li B 

PROVIDER: S-EPMC6065360 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature