An effective strategy to eliminate inherent cross-contamination in mtDNA next-generation sequencing of multiple samples

In this study, a novel sequencing strategy based on a unique double-barcode design was established. The results showed that when single barcode-based analysis strategy was used, cross-contamination level of 20 DNA samples ranged from 0.27% to 11.90% on Hiseq 2500 and from 0.93% to 17.70% on Hiseq X ten, whereas double barcode–based strategy could effectively eliminate cross-contamination. Moreover, the data indicated that cross-contamination was mainly derived from capture process and significantly affected by different NGS platforms. In addition, contamination level was negatively related to sequencing depth. Moreover, cross-contamination significantly increased the false positive calling of mtDNA heteroplasmic mutations and remarkably affected the heteroplasmy level of mtDNA mutations. In contrast, cross-contamination had no notable effect on classification of mtDNA haplogroup. Taken together, our novel double barcode–based sequencing strategy is very effective in eliminating cross-contamination, enhancing the detection accuracy of mtDNA NGS and improving its application in diagnosis or monitoring of diseases associated with mtDNA mutations.
Source: The Journal of Molecular Diagnostics - Category: Pathology Source Type: research

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Source: Stem Cell Research - Category: Stem Cells Source Type: research
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Source: Fight Aging! - Category: Research Authors: Tags: Newsletters Source Type: blogs
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Source: Yakugaku Zasshi : Journal of the Pharmaceutical Society of Japan - Category: Drugs & Pharmacology Authors: Tags: Yakugaku Zasshi Source Type: research
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Source: Fight Aging! - Category: Research Authors: Tags: Newsletters Source Type: blogs
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