1. faecal Peritonitis: intraperitoneal injection of 200µl of a human faeces suspension (KBE 75 x 10^6/ml), 2. Sham controls: intraperitoneal injection of 200µl of physiological sodium chloride
Growth protocol
8 weeks old female C57/BL6 Jackson mice
Extracted molecule
total RNA
Extraction protocol
Preparation of total RNA from lung, liver and spleen. Tissue homogenization was conducted with the tissue lyser by Qiagen (Retsch, Haan, Germany) with a 5mm steel ball for 4 minutes at 30hz and processed for total RNA isolation as described by the manufacturer using RNeasy Mini Kit (Qiagen, Hilden, Germany). RNA-Isolation from whole blood was performed according to the protocol of PAXgene Blood RNA Kit (PreAnalytix®, Qiagen, Hilden, Germany). Additional clearing from globin RNA was used for RNA from whole blood samples according to the GLOBINclearTM Mouse/Rat Kit by Ambion® (Applied Biosystems, Darmstadt, Germany) RNA integrity and concentration analyses were performed with the Bioanalyzer 2100 (Agilent Technologies®, Palo Alto, CA, USA) according to the manufacturers protocol RNA 6000 Nano Assay using RNA 6000 Nano LabChip® Kits by Agilent Technologies®.
Label
biotin
Label protocol
Biotinylated cRNA were prepared with the Ambion MessageAmp kit for Illumina arrays
Hybridization protocol
For hybridisation, the Whole-Genome Gene Expression with IntelliHybTM Seal protocoll (Revision B) from Illumina® (San Diego, CA, USA) was used.
Scan protocol
Scanning of arrays was done with the BeadArray Reader 500 X from Illumina®, San Diego, CA, USA).
Description
4338498057_E
Data processing
Gene chips from 44 samples (four tissues; four replicates for 6 h and 24 h post-infection and three replicates sham) were scanned and pre-processed using BeadStudio (version 3.23). Data analysis was carried out using R statistics software (version 2.92) and BioConductor (version 2.3). A robust spline normalization, combining the features of quantile and loess normalization, was conducted for the tissue separated sample groups (liver, lung, spleen and blood) and log 2 signals were obtained.
