Yamamoto et al. BMC Genomics 2012, 13:335 http://www.biomedcentral.com/1471-2164/13/335 Table 3 Top 25 GO categories overrepresented in SPL of 5WexGF mice compared with GF mice GOID GO:0007507 GO:0006952 GO:0016192 GO:0015992 GO:0001764 GO:0007283 GO:0006816 GO:0030036 GO:0042127 GO:0016055 GO:0007010 GO:0016337 GO:0018108 GO:0008610 GO:0009987 GO:0006954 GO:0006874 GO:0007166 GO:0005975 GO:0016311 GO:0042981 GO:0007420 GO:0006935 GO:0001701 GO:0009408 Term heart development defense response vesicle-mediated transport proton transport neuron migration Spermatogenesis calcium ion transport actin cytoskeleton organization and biogenesis regulation of cell proliferation Wnt receptor signaling pathway cytoskeleton organization and biogenesis cell-cell adhesion peptidyl-tyrosine phosphorylation lipid biosynthetic process cellular process inflammatory response cellular calcium ion homeostasis cell surface receptor linked signal transduction carbohydrate metabolic process Dephosphorylation regulation of apoptosis brain development Chemotaxis in utero embryonic development response to heat Number of Probes 91 63 76 64 32 94 41 52 46 61 43 33 25 76 78 48 23 54 81 61 83 30 45 55 26 Page 13 of 15 p_int 1.98E-13 4.39E-12 3.47E-11 1.69E-09 2.46E-09 2.65E-09 3.52E-09 3.81E-09 8.17E-09 1.16E-08 1.52E-08 2.30E-08 2.51E-08 2.69E-08 3.52E-08 4.44E-08 6.93E-08 9.30E-08 1.01E-07 1.76E-07 2.21E-07 3.64E-07 4.40E-07 5.95E-07 8.93E-07 MGP analysis was performed on GO BP categories which contains 4 ~ 99 genes (“Number of Probes”). The categories were sorted according to the integrated p value (p_int) and top 25 are represented. Bioanalyzer (Agilent Biotechnologies, Boeblingen, Ger- many) and consistently demonstrated high-quality RNA (28 S/18 S ratio, ~2). The labeled cRNA prepared from 200 ng total RNA by in vitro transcription (Enzo Bio- chem, New York, NY) was fragmented, hybridized to a Mouse Expression 430 array (Affymetrix, Santa Clara, CA) using an Affymetrix fluidics station, and scanned with an Affymetrix scanner, according to the Affymetrix protocol. Data were uploaded to the Center for Infor- mation Biology Gene Expression Database (CIBEX, http://cibex.nig.ac.jp/index.jsp) and are available under accession ID CBX256. Data were analyzed using the Affymetrix Microarray Suite [37] v.5.0 with all of the parameters set at default values (a global normalization was applied). Probe sets that had 2 or 3 absent A MAS detection calls per group (3 samples) in all groups were excluded. Therefore, genes that had more than 2 present calls in any one of the groups were included in the analysis. Probe annotations were obtained from the Affymetrix NetAffx Analysis Center. Functional and sig- nal pathway annotation of transcripts was based on Gene Ontology [47] and TRANSPATH [48] term assignments, respectively. Data clustering Hierarchical clustering of subsets of genes was performed using clustering and analysis software (Cluster 3.0; http://bonsai.hgc.jp/~mdehoon/software/ cluster/software.htm). The Pearson correlation coeffi- cient (r) was chosen to compute distances between ex- pression vectors (d = 1-r), and the complete linkage clustering algorithm was used to build the hierarchical tree. MetaGene Profiler (MGP) MGP has been developed to evaluate the significance of predefined sets of genes from transcriptome data (http:// metagp.ism.ac.jp/) [34]. The method, which employs a