title: ISO_soil_bacteria_fungi_diversity_study
organism: soil metagenome
description: Five different soils representing forest, grassland and arable biomes were collected from across France. All necessary permits were obtained from the respective land owners (INRA, ADEME, and private owners) for the described field studies. Five individual cores (20 cm depth) were sampled at each site using an unaligned sampling design within a defined area. Replicate soil cores were then bulked to obtain a composite sample for each site. After sieving soil samples to <4 mm, aliquots of 50 g were stored at -40degC prior to DNA extraction. Several physico-chemical parameters were measured for each soil: texture, pH, CaCO3, and total C and N. Physical and chemical analyses were performed by the Soil Analysis Laboratory of INRA (Arras, France, http://www.lille.inra.fr/las) using standard procedures (Plassart et al., 2012 - PLOS ONE).
attributes.tag: Acronym of the soil name
attributes.value: Res
attributes.tag: Acronym of the used DNA extraction method
attributes.value: ISOm
attributes.tag: ENA first public
attributes.value: 2014-07-31
attributes.tag: ENA last update
attributes.value: 2016-10-21
attributes.tag: External Id
attributes.value: SAMEA2242489
attributes.tag: INSDC center alias
attributes.value: INRA-CMSE
attributes.tag: INSDC center name
attributes.value: UMR Agroecology, INRA - Burgundy University, Dijon, France
attributes.tag: INSDC first public
attributes.value: 2014-07-31T17:01:03Z
attributes.tag: INSDC last update
attributes.value: 2016-10-21T09:33:02Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Number of the biological replicate
attributes.value: 2
attributes.tag: Submitter Id
attributes.value: ISOm_Res_16S_02
attributes.tag: collection date
attributes.value: 2011
attributes.tag: current land use
attributes.value: Crop soil
attributes.tag: depth
attributes.value: 0.1
attributes.tag: environment (biome)
attributes.value: cropland biome
attributes.tag: environment (feature)
attributes.value: farm
attributes.tag: environment (material)
attributes.value: agricultural soil
attributes.tag: geographic location (countryand/orsea,region)
attributes.value: France
attributes.tag: geographic location (elevation)
attributes.value: 46
attributes.tag: geographic location (latitude)
attributes.value: 49.028516
attributes.tag: geographic location (longitude)
attributes.value: 2.175321
attributes.tag: investigation type
attributes.value: mimarks-survey
attributes.tag: multiplex identifiers
attributes.value: CGTGAGCTGA
attributes.tag: nucleic acid extraction
attributes.value: This protocol is a modified version of ISO-11063 standard standard which includes a different mechanical lysis step (FastPrep bead-beating instead of the recommended bead-beating). Soil was added to a 15 mL Falcon tube containing 2.5 g of 1.4 mm diameter ceramic beads, 2 g of 106 µm diameter silica beads and and four glass beads of 4 mm diameter. Each soil sample was mixed with a solution of 100 mM Tris-HCl (pH 8), 100 mM EDTA (pH8), 100 mM NaCl, 2% (w/v) polyvinylpyrrolidone (40 g/mol) and 2% (w/v) sodium dodecyl sulfate. Tubes were then shaken for 3x30s at 4 m/sec in a FastPrep-24 (MP-Biomedicals, NY, USA) before incubation for 10 min at 70degC and centrifugation at 14,000g for 1 min. After removing the supernatant, proteins were precipitated with 1/10 volume of 3 M sodium acetate prior to centrifugation (14,000g for 5 min at 4degC). Finally, nucleic acids were precipitated by adding 1 volume of ice-cold isopropanol. The DNA pellets obtained after centrifugation (14,000g for 5 min at 4degC) were washed with 70% ethanol.
attributes.tag: pcr conditions
attributes.value: 5ng of DNA were used for a 25µl PCR conducted under the following conditions: initial denaturation: 94degC_2.0min; 35 cycles of denaturation: 94degC_0.5min;annealing: 52degC_0.5min;elongation: 72degC_1.0min;final elongation: 72degC_7.0min. A second PCR of 9 cycles was then conducted under similar PCR conditions with purified PCR products and ten base pair multiplex identifiers added to the primers at 5' position to specifically identify each sample and avoid PCR biases.
attributes.tag: pcr primers
attributes.value: F479:5-CAGCMGCYGCNGTAANAC-3; R888:5-CCGYCAATTCMTTTRAGT-3
attributes.tag: ph
attributes.value: 7.5
attributes.tag: project name
attributes.value: Ecofinders
attributes.tag: sample name
attributes.value: ISOm_Res_16S_02
attributes.tag: sample weight for DNA extraction
attributes.value: 1
attributes.tag: sequencing method
attributes.value: 454-Pyrosequencing
attributes.tag: soil environmental package
attributes.value: soil
attributes.tag: target gene
attributes.value: 16S rDNA
attributes.tag: texture
attributes.value: 81/11/8
attributes.tag: total n
attributes.value: 2.2
attributes.tag: total organic carbon
attributes.value: 50.2