Comparative analysis of ForenSeq mtDNA Control Region Kit, precision ID mtDNA Control Region Panel and Sanger...

DNA analysis in forensic science involves short tandem repeat analysis using genomic DNA, however, it is limited by low DNA concentration. Then, mitochondrial DNA (mtDNA) can be used as an alternative. In this study, to analyze mtDNA control region, we conducted an analysis using Sanger sequencing and two NGS methods: ForenSeq mtDNA Control Region Kit and Precision ID mtDNA Control Region Panel. This study is a comparative analysis of two commercially available NGS kits for mtDNA control region analysis. MtDNA was extracted from 75 Korean samples, and sequenced using Sanger sequencing, ForenSeq mtDNA Control Region Kit, or Precision ID mtDNA Control Region Panel. Although most mtDNA control region showed identical results between Sanger sequencing and NGS, differences in particular regions were observed in 43 of 75 samples using Sanger sequencing, ForenSeq mtDNA Control Region Kit, and Precision ID mtDNA Control Region Panel. These sites were poly-C sites of 16184-16193 in HV1, 303-315 in HV2, and 568-573 in HV3 that have been observed in InDel or substitution variant frequently. Additionally, heteroplasmy that was not observed in Sanger sequencing was detected in 6 samples in the NGS analysis. Therefore, mtDNA analysis using NGS is a convenient method for analyzing a large number of samples at once and is advantageous for detecting heteroplasmy, however, in length-based variation region, as bias can occur due to potential sequencing or alignment artifacts, caution should be taken in the interpretation of repetitive regions.