Inter-laboratory evaluation of the ISO standard 11063 "Soil quality - Method to directly extract DNA from soil samples" |
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Authors: | Petric I Philippot L Abbate C Bispo A Chesnot T Hallin S Laval K Lebeau T Lemanceau P Leyval C Lindström K Pandard P Romero E Sarr A Schloter M Simonet P Smalla K Wilke B-M Martin-Laurent F |
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Affiliation: | a Wellience Agro-Environment, BP 66517, 21065 Dijon Cedex, Franceb INRA, Université de Bourgogne, Soil and Environmental Microbiology, 21065 Dijon Cedex, Francec DACPA, Sezione di Scienze Agrochimiche, University of Catania, 95123 Catania, Italyd ADEME, Agriculture and Forestry Department, F-49004 Angers Cedex 01, Francee Laboratoire Etudes et Expertises, IPL santé, environnement durables Est, 54521 Maxéville, Francef Swedish University of Agricultural Sciences, Department of Microbiology, Box 7025, 750 07 Uppsala, Swedeng Laboratoire BioSol, APCA/Ecole d'Ingénieurs en Agriculture, 76000 Rouen, Franceh Plate-Forme Technologique AGROSYSTEMES, Institut Universitaire de Technologie, BP 50568, 68008 Colmar Cedex, Francei LIMOS, Nancy Université, CNRS, Faculté des Sciences, BP 239, 54506 Vandoeuvre-les-Nancy-Cedex, Francej Department of Food and Environmental Sciences, P.O. Box 56, 00014, University of Helsinki, Finlandk INERIS, Parc Technologique ALATA, BP 2, 60550 Verneuil en Halatte, Francel CSIC, Estacion Experimental del Zaidín, Environmental Protection, 18008 Granada, Spainm Department for Terrestrial Ecogenetics Helmholtz Zentrum München, German Research Center for Environmental Health (GmbH), D-85764 Neuherberg, Germanyn Environmental Microbial Genomics Group, Laboratoire AMPERE, Ecole Centrale de Lyon, Université de Lyon, 69134 Ecully, Franceo Julius Kühn-Institut Bundesforschungsinstitut für Kulturpflanzen, Institut für Epidemiologie und Pathogendiagnostik, D-38104 Braunschweig, Germanyp Technische Universität Berlin, Department of Ecology, D-10587 Berlin, Germany |
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Abstract: | Extracting DNA directly from micro-organisms living in soil is a crucial step for the molecular analysis of soil microbial communities. However, the use of a plethora of different soil DNA extraction protocols, each with its own bias, makes accurate data comparison difficult. To overcome this problem, a method for soil DNA extraction was proposed to the International Organization for Standardization (ISO) in 2006. This method was evaluated by 13 independent European laboratories actively participating in national and international ring tests. The reproducibility of the standardized method for molecular analyses was evaluated by comparing the amount of DNA extracted, as well as the abundance and genetic structure of the total bacterial community in the DNA extracted from 12 different soils by the 13 laboratories. High quality DNA was successfully extracted from all 12 soils, despite different physical and chemical characteristics and a range of origins from arable soils, through forests to industrial sites. Quantification of the 16S rRNA gene abundances by real time PCR and analysis of the total bacterial community structure by automated ribosomal intergenic spacer analysis (A-RISA) showed acceptable to good levels of reproducibility. Based on the results of both ring-tests, the method was unanimously approved by the ISO as an international standard method and the normative protocol will now be disseminated within the scientific community. Standardization of a soil DNA extraction method will improve data comparison, facilitating our understanding of soil microbial diversity and soil quality monitoring. |
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Keywords: | Soil DNA extraction Standardization Inter-laboratory assay DNA fingerprint qPCR |
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