Optimising the detection of marine taxonomic richness using environmental DNA metabarcoding: the effects of filter material, pore size and extraction method
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In: Metabarcoding and Metagenomics, Vol. 2, e28963, 02.11.2018.
Research output: Contribution to journal › Article › peer-review
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TY - JOUR
T1 - Optimising the detection of marine taxonomic richness using environmental DNA metabarcoding: the effects of filter material, pore size and extraction method
AU - Deiner, Kristy
AU - Lopez, Jacqueline
AU - Bourne, Steve
AU - Holman, Luke E.
AU - Seymour, Mathew
AU - Grey, Erin K.
AU - Lacoursiere-Roussel, Anais
AU - Li, Yiyuan
AU - Renshaw, Mark A.
AU - Pfrender, Michael E.
AU - Rius, Marc
AU - Bernatchez, Louis
AU - Lodge, David M.
PY - 2018/11/2
Y1 - 2018/11/2
N2 - The analysis of environmental DNA (eDNA) using metabarcoding has increased in use as a method for tracking biodiversity of ecosystems. Little is known about eDNA in marine human-modified environments, such as commercial ports, which are key sites to monitor for anthropogenic impacts on coastal ecosystems. To optimise an eDNA metabarcoding protocol in these environments, seawater samples were collected in a commercial port and methodologies for concentrating and purifying eDNA were tested for their effect on eukaryotic DNA yield and subsequent richness of Operational Taxonomic Units (OTUs). Different filter materials [Cellulose Nitrate (CN) and Glass Fibre (GF)], with different pore sizes (0.5 μm, 0.7 μm and 1.2 μm) and three previously published liquid phase extraction methods were tested. The number of eukaryotic OTUs detected differed by a factor of three amongst the method combinations. The combination of CN filters with phenol-chloroform-isoamyl alcohol extractions recovered a higher amount of eukaryotic DNA and OTUs compared to GF filters and the chloroform-isoamyl alcohol extraction method. Pore size was not independent of filter material but did affect the yield of eukaryotic DNA. For the OTUs assigned to a highly successful non-indigenous species, Styela clava, the two extraction methods with phenol significantly outperformed the extraction method without phenol; other experimental treatments did not contribute significantly to detection. These results highlight that careful consideration of methods is warranted because choice of filter material and extraction method create false negative detections of marine eukaryotic OTUs and underestimate taxonomic richness from environmental samples
AB - The analysis of environmental DNA (eDNA) using metabarcoding has increased in use as a method for tracking biodiversity of ecosystems. Little is known about eDNA in marine human-modified environments, such as commercial ports, which are key sites to monitor for anthropogenic impacts on coastal ecosystems. To optimise an eDNA metabarcoding protocol in these environments, seawater samples were collected in a commercial port and methodologies for concentrating and purifying eDNA were tested for their effect on eukaryotic DNA yield and subsequent richness of Operational Taxonomic Units (OTUs). Different filter materials [Cellulose Nitrate (CN) and Glass Fibre (GF)], with different pore sizes (0.5 μm, 0.7 μm and 1.2 μm) and three previously published liquid phase extraction methods were tested. The number of eukaryotic OTUs detected differed by a factor of three amongst the method combinations. The combination of CN filters with phenol-chloroform-isoamyl alcohol extractions recovered a higher amount of eukaryotic DNA and OTUs compared to GF filters and the chloroform-isoamyl alcohol extraction method. Pore size was not independent of filter material but did affect the yield of eukaryotic DNA. For the OTUs assigned to a highly successful non-indigenous species, Styela clava, the two extraction methods with phenol significantly outperformed the extraction method without phenol; other experimental treatments did not contribute significantly to detection. These results highlight that careful consideration of methods is warranted because choice of filter material and extraction method create false negative detections of marine eukaryotic OTUs and underestimate taxonomic richness from environmental samples
KW - eDNA
KW - 18SS ribsomal
KW - Seawater
KW - High-throughout sequencing
KW - Metazoaneukaryotes
KW - Non-indigenous species
U2 - 10.3897/mbmg.2.28963
DO - 10.3897/mbmg.2.28963
M3 - Article
VL - 2
JO - Metabarcoding and Metagenomics
JF - Metabarcoding and Metagenomics
SN - 2534-9708
M1 - e28963
ER -