Dataset Information

Metabarcoding of marine environmental DNA based on mitochondrial and nuclear genes.

ABSTRACT: We establish the new approach of environmental DNA (eDNA) analyses for the North Sea. Our study uses a multigene approach, including the mitochondrial cytochrome-c-oxidase subunit I (COI) gene for analyzing species composition and the nuclear hypervariable region V8 of 18S rDNA for analyzing supraspecific biodiversity. A new minibarcode primer (124?bp) was created on the basis of a metazoan COI barcode library with 506 species and tested in silico, in vitro, and in situ. We applied high throughput sequencing to filtrates of 23 near-bottom water samples taken at three seasons from 14 stations. The set of COI primers allowed amplification of mitochondrial minibarcodes for diverse metazoan phyla and the differentiation at the species level for more than 99% of the specimens in the dataset. Our results revealed that the number of sequences is not consistent with proportions in the given DNA mixture. Altogether, environmental sequences could be assigned to 114 species and to 12 metazoan phyla. A spatial distribution of taxa recovered by eDNA was congruent with known distributions. Finally, the successful detection of species and biodiversity depends on a comprehensive sequence reference database. Our study offers a powerful tool for future biodiversity research, including the detection of nonnative species.

SUBMITTER: Gunther B

PROVIDER: S-EPMC6172225 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Metabarcoding of marine environmental DNA based on mitochondrial and nuclear genes.

Günther Babett B Knebelsberger Thomas T Neumann Hermann H Laakmann Silke S Martínez Arbizu Pedro P

Scientific reports 20181004 1

We establish the new approach of environmental DNA (eDNA) analyses for the North Sea. Our study uses a multigene approach, including the mitochondrial cytochrome-c-oxidase subunit I (COI) gene for analyzing species composition and the nuclear hypervariable region V8 of 18S rDNA for analyzing supraspecific biodiversity. A new minibarcode primer (124 bp) was created on the basis of a metazoan COI barcode library with 506 species and tested in silico, in vitro, and in situ. We applied high throughp ...[more]

PMID: 30287908

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Project description:DNA extraction from environmental samples (environmental DNA; eDNA) for metabarcoding-based biodiversity studies is gaining popularity as a noninvasive, time-efficient, and cost-effective monitoring tool. The potential benefits are promising for marine conservation, as the marine biome is frequently under-surveyed due to its inaccessibility and the consequent high costs involved. With increasing numbers of eDNA-related publications have come a wide array of capture and extraction methods. Without visual species confirmation, inconsistent use of laboratory protocols hinders comparability between studies because the efficiency of target DNA isolation may vary. We determined an optimal protocol (capture and extraction) for marine eDNA research based on total DNA yield measurements by comparing commonly employed methods of seawater filtering and DNA isolation. We compared metabarcoding results of both targeted (small taxonomic group with species-level assignment) and universal (broad taxonomic group with genus/family-level assignment) approaches obtained from replicates treated with the optimal and a low-performance capture and extraction protocol to determine the impact of protocol choice and DNA yield on biodiversity detection. Filtration through cellulose-nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit outperformed other combinations of capture and extraction methods, showing a ninefold improvement in DNA yield over the poorest performing methods. Use of optimized protocols resulted in a significant increase in OTU and species richness for targeted metabarcoding assays. However, changing protocols made little difference to the OTU and taxon richness obtained using universal metabarcoding assays. Our results demonstrate an increased risk of false-negative species detection for targeted eDNA approaches when protocols with poor DNA isolation efficacy are employed. Appropriate optimization is therefore essential for eDNA monitoring to remain a powerful, efficient, and relatively cheap method for biodiversity assessments. For seawater, we advocate filtration through cellulose-nitrate membranes and extraction with Qiagen's DNeasy Blood & Tissue Kit or phenol-chloroform-isoamyl for successful implementation of eDNA multi-marker metabarcoding surveys.

Dataset Information

Metabarcoding of marine environmental DNA based on mitochondrial and nuclear genes.

Publications

Metabarcoding of marine environmental DNA based on mitochondrial and nuclear genes.

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