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Characterization and remediation of sample index swaps by non-redundant dual indexing on massively parallel sequencing platforms

By Maura Costello, Mark Fleharty, Justin Abreu, Yossi Farjoun, Steven Ferriera, Laurie Holmes, Tom Howd, Tamara Mason, Gina Vicente, Michael Dasilva, Wendy Brodeur, Timothy DeSmet, Sheila Dodge, Niall J. Lennon, Stacey Gabriel

Posted 10 Oct 2017
bioRxiv DOI: 10.1101/200790 (published DOI: 10.1186/s12864-018-4703-0)

Here, we present an in-depth characterization of the index swapping mechanism on Illumina instruments that employ the ExAmp chemistry for cluster generation (HiSeqX, HiSeq4000, and NovaSeq). We discuss best practices for eliminating the effects of index swapping on data integrity by utilizing unique dual indexing for complete filtering of index swapped reads. We calculate mean swap rates across multiple sample preparation methods and sequencer models, demonstrating that different methods can have vastly different swap rates, and show that even non-ExAmp chemistry instruments display trace levels of index swapping. Finally, using computational methods we provide a greater insight into the mechanism of index swapping.

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