Comparison of mtDNA sequence variations

We applied the 3–97% threshold to mtDNA sequences to identify homoplasmy and heteroplasmy between two DNA sources. That is, a site was identified as homoplasmy if AAF > 97% and a site was deemed as heteroplasmy if AAF was between 3 and 97%. We excluded any sites with coverage < 250-fold, and we also excluded the following artifact prone sites: 301, 302, 310, 316, 3107, 16,182, according to the guidance from the mtDNA calling pipeline by Broad Institute (gnomAD v3.1)³¹. We compared the distribution of homoplasmic and heteroplasmic sites between the paired samples. We also compared the distribution of AAFs for identified mtDNA variation sites between paired samples. We used MToolBox²⁷ to identify mtDNA sequence variations from whole blood-derived DNA samples. The identification of mtDNA sequence variations in LCL-derived DNA samples was previously described¹¹. All statistical analyses for comparisons used R language³².