Maternal transcripts are indispensable and play an essential role in regulation of the resumption of meiosis, nuclear reprogramming and subsequent development. In this study, we conducted interactive subtraction analysis to investigate the dynamic changes in the gene expression profiles of mouse oocytes after artificial activation. To accomplish this, we compared the cDNA libraries of freshly ovulated metaphase II (MII) oocytes and pronuclear-stage eggs at 8 h after activation, which were obtained by interactive subtraction. After constructing the cDNA libraries, we sequenced 635 subtracted clones of differentially expressed genes and the identities of 515 of these genes were determined by online BLAST analysis. The analysis revealed that genes down-regulated after activation were mainly responsible for enzyme and receptor activities, and also for DNA, chromatin, ATP, protein, and ion binding. On the other hand, genes up-regulated after activation were mainly responsible for enzyme activities, ion binding, and the maintenance of cell structure. For further insight into the functions of the genes, gene network analysis was performed, and in order to validate differential gene expression, the gene expression levels were determined by using real-time quantitative PCR. The present analysis was capable of identifying the genes that changed dynamically following the resumption of meiosis, and these genes may control the developmental programs in oocytes and preimplantation embryos.