Evaluation of the ISO Standard 11063 DNA extraction procedure for assessing soil microbial abundance and community structure
Plassart, Pierre; Terrat, Sebastien; Thomson, Bruce; Griffiths, Robert; Dequiedt, Samuel; Lelievre, Melanie; Regnier, Tiffanie; Nowak, Virginie; Bailey, Mark; Lemanceau, Philippe; Bispo, Antonio; Chabbi, Abad; Maron, Pierre-Alain; Mougel, Christophe; Ranjard, Lionel. 2012 Evaluation of the ISO Standard 11063 DNA extraction procedure for assessing soil microbial abundance and community structure. PLoS ONE, 7 (9), e44279. 8, pp. 10.1371/journal.pone.0044279Before downloading, please read NORA policies.
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Soil DNA extraction has become a critical step in describing microbial biodiversity. Historically, ascertaining overarching microbial ecological theories has been hindered as independent studies have used numerous custom and commercial DNA extraction procedures. For that reason, a standardized soil DNA extraction method (ISO-11063) was previously published. However, although this ISO method is suited for molecular tools such as quantitative PCR and community fingerprinting techniques, it has only been optimized for examining soil bacteria. Therefore, the aim of this study was to assess an appropriate soil DNA extraction procedure for examining bacterial, archaeal and fungal diversity in soils of contrasting land-use and physico-chemical properties. Three different procedures were tested: the ISO-11063 standard; a custom procedure (GnS-GII); and a modified ISO procedure (ISOm) which includes a different mechanical lysis step (a FastPrep ®-24 lysis step instead of the recommended bead-beating). The efficacy of each method was first assessed by estimating microbial biomass through total DNA quantification. Then, the abundances and community structure of bacteria, archaea and fungi were determined using real-time PCR and terminal restriction fragment length polymorphism approaches. Results showed that DNA yield was improved with the GnS-GII and ISOm procedures, and fungal community patterns were found to be strongly dependent on the extraction method. The main methodological factor responsible for differences between extraction procedure efficiencies was found to be the soil homogenization step. For integrative studies which aim to examine bacteria, archaea and fungi simultaneously, the ISOm procedure results in higher DNA recovery and better represents microbial communities.
|Item Type:||Publication - Article|
|Digital Object Identifier (DOI):||10.1371/journal.pone.0044279|
|Programmes:||CEH Topics & Objectives 2009 - 2012 > Biodiversity > BD Topic 1 - Observations, Patterns, and Predictions for Biodiversity > BD - 1.1 - Standards for data collection, quality, management and integration ...
CEH Topics & Objectives 2009 - 2012 > Biodiversity
|CEH Sections:||Directors, SPCs
|Additional Pages:||1-page supplementary figure|
|Additional Information. Not used in RCUK Gateway to Research.:||This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited|
|NORA Subject Terms:||Agriculture and Soil Science|
|Date made live:||13 Sep 2012 15:59|
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