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Biases in bulk: DNA metabarcoding of marine communities and the methodology involved
  • Luna van der Loos,
  • Reindert Nijland
Luna van der Loos
Wageningen Universiteit en Research

Corresponding Author:[email protected]

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Reindert Nijland
, Wageningen Universiteit en Research
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With the growing anthropogenic pressure on marine ecosystems, the need for efficient monitoring of biodiversity grows stronger. DNA metabarcoding of bulk samples is increasingly implemented in ecosystem assessments and is more cost-efficient and less time-consuming than monitoring based on morphology. However, before raw sequences are obtained from bulk samples, a profound number of methodological choices must be made. Here, we critically review the recent methods used for metabarcoding of marine bulk samples (including benthic, plankton and diet samples) and indicate how potential biases can be introduced throughout sampling, pre-processing, DNA extraction, marker and primer selection, PCR amplification and sequencing. From a total of 64 studies evaluated, our recommendations for best practices include to (a) consider DESS as a fixative instead of ethanol, (b) use the DNeasy PowerSoil kit for any samples containing traces of sediment, (c) not limit the marker selection to COI only, but preferably include multiple markers for higher taxonomic resolution, (d) avoid touchdown PCR profiles, (e) use a fixed annealing temperature for each primer pair when comparing across studies or institutes, (f) use a minimum of 3 PCR replicates and (g) include both negative and positive controls. Although the implementation of DNA metabarcoding still faces several technical complexities, we foresee wide-ranging advances in the near future, including improved bioinformatics for taxonomic assignment, sequencing of longer fragments, and the use of whole-genome information. Despite the bulk of biases involved in metabarcoding of bulk samples, it is clear that DNA metabarcoding provides a valuable tool in ecosystem assessments.
26 Jun 2020Submitted to Molecular Ecology
11 Jul 2020Reviewer(s) Assigned
22 Jul 2020Review(s) Completed, Editorial Evaluation Pending
28 Jul 2020Editorial Decision: Accept
Jul 2021Published in Molecular Ecology volume 30 issue 13 on pages 3270-3288. 10.1111/mec.15592