Genechip analysis suite software

For genome-wide transcription analysis, the GeneChip Human Genome U133 Plus 2.0 array was used as previously described [49 (link), 50 (link)]. Briefly, total RNA was extracted with ISOGEN (Nippon Gene) from LNCaP, LNCaP-SF, and LNCaP-SF cells treated with siRNAs (non-targeting control, siAR-1, and siAR-2). Following in vitro transcription (IVT) and cRNA fragmentation, the fragmented IVT product was hybridized on an array and stained with streptavidin phycoerythrin according to the manufacturer's recommended protocol. The arrays were scanned using the Affymetrix GeneChip Scanner 3000 (Affymetrix), and GeneChip Analysis Suite software program version 5.0 (Affymetrix) was used to calculate the signal value for each gene probe.

Microarray and computational gene expression analyses were performed using a GeneChip® system with a Human Genome U133-plus 2.0 array (Affymetrix, Santa Clara), which was spotted with approximately 54,000 probe sets, as previously described [9] (link), [10] (link), [11] (link), [12] (link). Samples for array hybridization were prepared as described in the Affymetrix GeneChip® Expression Technical Manual. The scanned arrays were analyzed using the GeneChip Analysis Suite Software (Affymetrix). The obtained hybridization intensity data and qualities were checked using the GeneSpring® software.

Transcriptome analysis was performed on mouse colon samples as described previously (Nagata et al., 2017 (link)). In brief, total RNA was extracted from the distal colons on day 7 after initiation of DSS treatment. mRNA was purified from the mixture of total RNA using an RNeasy Mini Kit (Qiagen, Crawley, United Kingdom), and mRNA was pooled from three to six mice per group (WT normal, WT colitis and IL-4Rα-/- colitis). Microarray analysis was performed using a GeneChip Mouse Gene 1.0 ST Array (Affymetrix, Santa Clara, CA, United States). The GeneChip was scanned with a GeneChip Scanner 3000 (Affymetrix), and gene expression was analyzed using GeneChip Analysis Suite Software (Affymetrix). Median centered and log2 transformed transcript levels are indicated by the following color code: blue, low; red, high. Furthermore, the data were analyzed GeneChip microarray data using Transcriptome Analysis Console (TAC; Thermo Fisher Scientific, Waltham, MA, United States), GeneSpring (Silicon Genetics, Redwood City, CA, United States) and Ingenuity Pathway Analysis (IPA; Ingenuity Systems, Redwood City, CA, United States, http://www.ingenuity.com) to extract significant genes, and identify the gene ontology and the canonical pathways associated with the differentially expressed genes.

Gene expression was analysed using a GeneChip system with a Human Genome U133-plus 2.0 array, which was spotted with ~54 000 probe sets (Affymetrix Inc., Santa Clara, CA, USA). Samples for array hybridisation were prepared as described in the Affymetrix GeneChip Expression Technical Manual (Affymetrix Inc.). The scanned arrays were analysed using the GeneChip Analysis Suite Software (Affymetrix Inc.). The obtained hybridisation intensity data were analysed using the GeneSpring analysis software (Silicon Genetics, Redwood City, CA, USA) to extract the significant genes. To examine gene ontology, including the biological processes, cellular components, molecular functions, and gene networks, the obtained data were analysed using the Ingenuity Pathways Analysis tools (Ingenuity Systems Inc.), a web-delivered application that enables the identification, visualisation, and exploration of molecular interaction networks in gene-expression data.