Mouse Whole Transcriptome Expression Profiling

Per offspring, colon and SI samples were analysed as described previously [43 (link)], and in total, six male and six female samples were used in this analysis (three of the nine females were not included in the microarray analysis due to budget limitations). In brief, 100 ng of purified RNA was used for the preparation of labelled cDNA, applying the Ambion Whole Transcript (WT) Expression Kit (Life Technologies, Carlsbad, CA, USA) in combination with the Affymetrix GeneChip WT Terminal Labeling Kit (Affymetrix, Santa Clara, CA, USA). All samples were hybridized at one time point to Affymetrix GeneChip Mouse Gene 1.1 ST arrays according to standard Affymetrix protocols. Quality control and normalization were performed using Bioconductor software packages integrated in an on-line pipeline [44 (link)]. Normalized expression estimates of probe sets were computed by the robust multiarray (RMA) analysis algorithm available in the Bioconductor library AffyPLM using default settings [45 (link)]. Probe sets were redefined according to Dai et al. [46 (link)] and assigned to unique gene identifiers (IDs) of the Entrez Gene database, resulting in 21,187 assigned Entrez IDs. Array data were submitted to the Gene Expression Omnibus and are available under accession number GSE57516.