There is increasing evidence to show that 2-cell stage mouse blastomeres have differing developmental properties. Additionally, it has been suggested that such a difference might be due to their distribution of mRNA and/or protein asymmetry. However, to date, the exact genes that are involved in the orientation and order of blastomere division are not known. In this study, some differentially expressed transcripts were identified. Axin1, cell division cycle 25 homolog C (Cdc25c) and cyclin-dependent inhibitor 2D (Cdkn2d) were selected for validation by real-time polymerase chain reaction (PCR) based on published data. Our real-time PCR results demonstrated that Axin1, Cdc25c and Cdkn2d genes had different levels of expression among blastomeres of the mouse 2-cell embryo i.e. the level of Axin1 mRNA was significantly higher in one blastomere when compared with the other blastomeres of the 2-cell embryo (p < 0.05). The variation in Cdc25c (p < 0.05) and Cdkn2d (p < 0.01) mRNA expression followed a similar trend to that of Axin1. In addition, the highest levels of expression of these three genes were detected in the same blastomere in the 2-cell embryo. We confirmed that there was an asymmetrical distribution pattern for Axin1, Cdc25c and Cdkn2d transcripts in 2-cell embryos. In conclusion, this study demonstrated clearly that there is embryonic asymmetry at the 2-cell stage and that these differentially expressed genes may result in differentiation in expression in embryo development.