Fungal community analysis by high-throughput sequencing of amplified markers – a user's guide

TY - JOUR

T1 - Fungal community analysis by high-throughput sequencing of amplified markers – a user's guide

AU - Lindahl, Björn D.

AU - Nilsson, R. Henrik

AU - Tedersoo, Leho

AU - Abarenkov, Kessy

AU - Carlsen, Tor

AU - Kjøller, Rasmus

AU - Kõljalg, Urmas

AU - Pennanen, Taina

AU - Rosendahl, Søren

AU - Stenlid, Jan

AU - Kauserud, Håvard

PY - 2013/7

Y1 - 2013/7

N2 - Novel high-throughput sequencing methods outperform earlier approaches in terms of resolution and magnitude. They enable identification and relative quantification of community members and offer new insights into fungal community ecology. These methods are currently taking over as the primary tool to assess fungal communities of plant-associated endophytes, pathogens, and mycorrhizal symbionts, as well as free-living saprotrophs. Taking advantage of the collective experience of six research groups, we here review the different stages involved in fungal community analysis, from field sampling via laboratory procedures to bioinformatics and data interpretation. We discuss potential pitfalls, alternatives, and solutions. Highlighted topics are challenges involved in: obtaining representative DNA/RNA samples and replicates that encompass the targeted variation in community composition, selection of marker regions and primers, options for amplification and multiplexing, handling of sequencing errors, and taxonomic identification. Without awareness of methodological biases, limitations of markers, and bioinformatics challenges, large-scale sequencing projects risk yielding artificial results and misleading conclusions.

AB - Novel high-throughput sequencing methods outperform earlier approaches in terms of resolution and magnitude. They enable identification and relative quantification of community members and offer new insights into fungal community ecology. These methods are currently taking over as the primary tool to assess fungal communities of plant-associated endophytes, pathogens, and mycorrhizal symbionts, as well as free-living saprotrophs. Taking advantage of the collective experience of six research groups, we here review the different stages involved in fungal community analysis, from field sampling via laboratory procedures to bioinformatics and data interpretation. We discuss potential pitfalls, alternatives, and solutions. Highlighted topics are challenges involved in: obtaining representative DNA/RNA samples and replicates that encompass the targeted variation in community composition, selection of marker regions and primers, options for amplification and multiplexing, handling of sequencing errors, and taxonomic identification. Without awareness of methodological biases, limitations of markers, and bioinformatics challenges, large-scale sequencing projects risk yielding artificial results and misleading conclusions.

U2 - 10.1111/nph.12243

DO - 10.1111/nph.12243

M3 - Journal article

C2 - 23534863

SN - 1469-8137

VL - 199

SP - 288

EP - 299

JO - New Phytologist

JF - New Phytologist

IS - 1

ER -