From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel

Despite a multitude of methods for the sample preparation, sequencing, and data analysis of mitochondrial DNA (mtDNA), the demand for innovation remains, particularly in comparison with nuclear DNA (nDNA) research. The Applied Biosystems™ Precision ID mtDNA Whole Genome Panel (Thermo Fisher Scientif...

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Autores principales:	Filipe Cortes-Figueiredo, Filipa S. Carvalho, Ana Catarina Fonseca, Friedemann Paul, José M. Ferro, Sebastian Schönherr, Hansi Weissensteiner, Vanessa A. Morais
Formato:	article
Lenguaje:	EN
Publicado:	MDPI AG 2021
Materias:	mitochondrial DNA next-generation sequencing massively parallel sequencing whole genome sequencing Precision ID Thermo Fisher Scientific Biology (General) QH301-705.5 Chemistry QD1-999
Acceso en línea:	https://doaj.org/article/0be1cdfc7bdb46ac8f91e4007aba17c7
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spelling	oai:doaj.org-article:0be1cdfc7bdb46ac8f91e4007aba17c72021-11-11T17:25:52ZFrom Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel10.3390/ijms2221120311422-00671661-6596https://doaj.org/article/0be1cdfc7bdb46ac8f91e4007aba17c72021-11-01T00:00:00Zhttps://www.mdpi.com/1422-0067/22/21/12031https://doaj.org/toc/1661-6596https://doaj.org/toc/1422-0067Despite a multitude of methods for the sample preparation, sequencing, and data analysis of mitochondrial DNA (mtDNA), the demand for innovation remains, particularly in comparison with nuclear DNA (nDNA) research. The Applied Biosystems™ Precision ID mtDNA Whole Genome Panel (Thermo Fisher Scientific, USA) is an innovative library preparation kit suitable for degraded samples and low DNA input. However, its bioinformatic processing occurs in the enterprise Ion Torrent Suite™ Software (TSS), yielding BAM files aligned to an unorthodox version of the revised Cambridge Reference Sequence (rCRS), with a heteroplasmy threshold level of 10%. Here, we present an alternative customizable pipeline, the PrecisionCallerPipeline (PCP), for processing samples with the correct rCRS output after Ion Torrent sequencing with the Precision ID library kit. Using 18 samples (3 original samples and 15 mixtures) derived from the 1000 Genomes Project, we achieved overall improved performance metrics in comparison with the proprietary TSS, with optimal performance at a 2.5% heteroplasmy threshold. We further validated our findings with 50 samples from an ongoing independent cohort of stroke patients, with PCP finding 98.31% of TSS’s variants (TSS found 57.92% of PCP’s variants), with a significant correlation between the variant levels of variants found with both pipelines.Filipe Cortes-FigueiredoFilipa S. CarvalhoAna Catarina FonsecaFriedemann PaulJosé M. FerroSebastian SchönherrHansi WeissensteinerVanessa A. MoraisMDPI AGarticlemitochondrial DNAnext-generation sequencingmassively parallel sequencingwhole genome sequencingPrecision IDThermo Fisher ScientificBiology (General)QH301-705.5ChemistryQD1-999ENInternational Journal of Molecular Sciences, Vol 22, Iss 12031, p 12031 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	mitochondrial DNA next-generation sequencing massively parallel sequencing whole genome sequencing Precision ID Thermo Fisher Scientific Biology (General) QH301-705.5 Chemistry QD1-999
spellingShingle	mitochondrial DNA next-generation sequencing massively parallel sequencing whole genome sequencing Precision ID Thermo Fisher Scientific Biology (General) QH301-705.5 Chemistry QD1-999 Filipe Cortes-Figueiredo Filipa S. Carvalho Ana Catarina Fonseca Friedemann Paul José M. Ferro Sebastian Schönherr Hansi Weissensteiner Vanessa A. Morais From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel
description	Despite a multitude of methods for the sample preparation, sequencing, and data analysis of mitochondrial DNA (mtDNA), the demand for innovation remains, particularly in comparison with nuclear DNA (nDNA) research. The Applied Biosystems™ Precision ID mtDNA Whole Genome Panel (Thermo Fisher Scientific, USA) is an innovative library preparation kit suitable for degraded samples and low DNA input. However, its bioinformatic processing occurs in the enterprise Ion Torrent Suite™ Software (TSS), yielding BAM files aligned to an unorthodox version of the revised Cambridge Reference Sequence (rCRS), with a heteroplasmy threshold level of 10%. Here, we present an alternative customizable pipeline, the PrecisionCallerPipeline (PCP), for processing samples with the correct rCRS output after Ion Torrent sequencing with the Precision ID library kit. Using 18 samples (3 original samples and 15 mixtures) derived from the 1000 Genomes Project, we achieved overall improved performance metrics in comparison with the proprietary TSS, with optimal performance at a 2.5% heteroplasmy threshold. We further validated our findings with 50 samples from an ongoing independent cohort of stroke patients, with PCP finding 98.31% of TSS’s variants (TSS found 57.92% of PCP’s variants), with a significant correlation between the variant levels of variants found with both pipelines.
format	article
author	Filipe Cortes-Figueiredo Filipa S. Carvalho Ana Catarina Fonseca Friedemann Paul José M. Ferro Sebastian Schönherr Hansi Weissensteiner Vanessa A. Morais
author_facet	Filipe Cortes-Figueiredo Filipa S. Carvalho Ana Catarina Fonseca Friedemann Paul José M. Ferro Sebastian Schönherr Hansi Weissensteiner Vanessa A. Morais
author_sort	Filipe Cortes-Figueiredo
title	From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel
title_short	From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel
title_full	From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel
title_fullStr	From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel
title_full_unstemmed	From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel
title_sort	from forensics to clinical research: expanding the variant calling pipeline for the precision id mtdna whole genome panel
publisher	MDPI AG
publishDate	2021
url	https://doaj.org/article/0be1cdfc7bdb46ac8f91e4007aba17c7
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From Forensics to Clinical Research: Expanding the Variant Calling Pipeline for the Precision ID mtDNA Whole Genome Panel

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