Regassa et al. BMC Genomics 2011, 12:57 http://www.biomedcentral.com/1471-2164/12/57 Page 8 of 20 Figure 5 The top significantly changed GO terms (biological processes) with the proportion of transcripts involved among those over expressed in GV oocytes and CCs. The proportion of transcripts in a GO term was calculated as the number of genes over expressed in one sample divided by the total number of genes that are involved in that given GO term multiplied by 100 (P < 0.001). carbohydrate metabolism while Figure 11 shows a network that plays a role in cellular development. Global transcriptome changes in the CCs and associated functional changes during COCs in vitro maturation In this experiment we analyzed global transcriptome changes during the transition of CCs from GV (number Figure 6 Hierarchical clustering and heat map of the top differentially expressed genes between oocytes cultured with (red bar) or without (green bar) their companion CCs with a fold change of more than 4. Abbreviations, OO + CCs and OO - CCs stand for oocytes cultured with or without their companion CCs, respectively and numbers (1, 2 and 3) indicate the three biological replicates that were used for microarray hybridization. Figure 7 Hierarchical clustering and heat map of the top differentially expressed genes between CCs cultured with (red bar) or without (green bar) their enclosed oocytes with a fold change of more than 16. Abbreviations, CCs + OO and CCs - OO stand for cumulus cells cultured with or without ooplasm, respectively. Numbers (1, 2 and 3) indicate the three biological replicates that were used for microarray hybridization. 9) to MII (number 10) stage. The results of MAS 5 pre- sent and absent call showed that of 12827 detected genes 4689 and 834 are exclusively expressed in GV and MII CCs, respectively, while 7304 are expressed com- monly at both stages (Additional files, 16, 17 and 18). Additionally, expression analysis of these detected genes showed that a total of 4677 genes are differentially expressed between the two samples of which 2397 and 2280 are over expressed in GV and MII stages, respec- tively (Additional file 19). The heat map and hierarchical clustering of the top differentially expressed genes (fold change > 256), GO categories (biological processes and cellular and molecular functions) between the two CCs stages are shown in Figures 12, 13 and 14, respectively. Validation of the microarray data using quantitative and semi quantitative real time RT-PCR In order to validate the micro array data, a total of 23 transcripts were quantified using qRT-PCR as described in materials and methods section. The assayed genes are selected based on the criteria of being abundant at sig- nificantly higher level or their exclusive abundance in either of the samples considered in the array analysis. Transcripts Semi-quantitative RT-PCR was also adopted to validate some genes that are exclusively expressed either as the oocyte or CC transcripts. With the