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A simple centrifugation protocol leads to a 55-fold mitochondrial DNA enrichment and paves the way for future mitogenomic research

Jan Niklas Macher, Vera Zizka, Alexander Martin Weigand, Florian Leese
doi: https://doi.org/10.1101/106583
Jan Niklas Macher
aAquatic Ecosystem Research, Faculty of Biology, University of Duisburg-Essen, Universitätsstraβe 5, 45141 Essen, Germany
bCentre for Water and Environmental Research, University of Duisburg-Essen, Universitätsstraβe 2, 45141 Essen, Germany
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  • For correspondence: jan.macher@uni-due.de
Vera Zizka
aAquatic Ecosystem Research, Faculty of Biology, University of Duisburg-Essen, Universitätsstraβe 5, 45141 Essen, Germany
bCentre for Water and Environmental Research, University of Duisburg-Essen, Universitätsstraβe 2, 45141 Essen, Germany
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Alexander Martin Weigand
aAquatic Ecosystem Research, Faculty of Biology, University of Duisburg-Essen, Universitätsstraβe 5, 45141 Essen, Germany
bCentre for Water and Environmental Research, University of Duisburg-Essen, Universitätsstraβe 2, 45141 Essen, Germany
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Florian Leese
aAquatic Ecosystem Research, Faculty of Biology, University of Duisburg-Essen, Universitätsstraβe 5, 45141 Essen, Germany
bCentre for Water and Environmental Research, University of Duisburg-Essen, Universitätsstraβe 2, 45141 Essen, Germany
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Abstract

DNA (meta)barcoding is used to study biodiversity and is available for standardised assessments. However, it suffers from PCR bias, which can lead to the loss of specific taxa. PCR-free techniques such as shotgun metagenomics are therefore thought to be more suited for biodiversity assessments, but are currently limited by incomplete reference libraries.

The technique of ‘mitogenome-skimming’ or ‘mitogenomics’, in which complete mitochondrial genomes are sequenced, is ideal to bridge the techniques of (meta)barcoding and metagenomics. However, without the enrichment of mitochondria, roughly 99 % of all sequencing reads are of non-mitochondrial origin and mostly useless for common applications, e.g. species identification.

Here, we present a simple centrifugation protocol that leads to an average 140-fold enrichment of mitochondrial DNA. By sequencing six ‘mock’- communities – comprising the freshwater taxa Corbicula fluminea, Gammarus roeselii and Hydropsyche exocellata each – we recovered whole mitochondrial genomes of these species and the acanthocephalan endoparasite Pomphorhynchus laevis.

The presented protocol will greatly speed up building reference libraries for whole mitochondrial genomes, as dozens of species could be sequenced on a single MiSeq run. Subsequently, it will also allow biodiversity assessments using mitogenomics at greatly reduced costs in comparison to mitogenomic approaches without prior enrichment for mitochondria.

Footnotes

  • JNM: jan.macher{at}uni-due.de, +49.201.183-6710>

  • VZ: vera.zizka{at}uni-due.de, +49.201.183-4053

  • AMW: alexander.weigand{at}uni-due.de, +49.201.183-6710

  • FL: florian.leese{at}uni-due.de, +49.201.183-4053

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
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Posted March 23, 2017.
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A simple centrifugation protocol leads to a 55-fold mitochondrial DNA enrichment and paves the way for future mitogenomic research
Jan Niklas Macher, Vera Zizka, Alexander Martin Weigand, Florian Leese
bioRxiv 106583; doi: https://doi.org/10.1101/106583
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A simple centrifugation protocol leads to a 55-fold mitochondrial DNA enrichment and paves the way for future mitogenomic research
Jan Niklas Macher, Vera Zizka, Alexander Martin Weigand, Florian Leese
bioRxiv 106583; doi: https://doi.org/10.1101/106583

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