Mouse clariom s mm10 microarray

Total RNA was extracted following the instructions of the manufacturer (TRIzol, Life Technologies) from 4T1 cells treated with and without valproic acid (VPA) at dose of 0.5 mM for 10 days. Microarray probes were synthetized in one cycle of RNA amplification in which molecules were labeled (Affymetrix microarray station, Affymetrix, CA). The labeled microarray probes were hybridized on a Mouse Clariom S (mm10) microarray (Thermo Fisher Scientific), and the CEL files of microarray data obtained from the Affymetrix platform were normalized using the RMA method in Affymetrix Expression Console software (Affymetrix, CA). Gene-set enrichment analysis was performed with the online java module of GSEA software, version 3.0, while a network-based gene-set enrichment analysis was performed with Cytoscape software, version 3.6.0. Bioinformatics analyses were performed in R version 3.4.1; the R-package made4 was used to create an expression heatmap using Euclidean distances and the Ward method, and the FactoMineR R-package was used to perform an unsupervised principal component analysis. Genes with significant differences in expression were selected with the SAM algorithm using a FDR threshold of 5 percent.

Total RNA was extracted from 4T1 tumors in treated and control mice, following the instructions of the manufacturer (TRIzol, Life Technologies). Microarray probes was synthetized by one cycle of RNA amplification in which molecules were labeled in an Affymetrix microarray station (Affymetrix, CA). Labeled microarray probes were hybridized on a Mouse Clariom S (mm10) microarray (Thermo Fisher Scientific). The CEL files of microarray data obtained from the Affymetrix platform were normalized using the RMA method in Affymetrix Expression Console software (Affymetrix, CA) (34 (link)). As described by Lyons et al. (35 (link)), gene modules of specific immune cell expression profiles were constructed for the purpose of immune-cell-specific profiling of cancer. With these specific immune modules, we conducted gene-set enrichment analyses using the online java module of GSEA software version 3.0 (36 (link)). A network-based analysis of gene enrichment was performed with Cytoscape software version 3.6.0 (31 (link)). Differentially expressed genes were identified with a rank products analysis. A functional analysis of genes that were upregulated in the vaccinated group was performed using the Gene Ontology biological process database and the DAVID application from the NIH website (37 (link)).