We concentrate on the critical distinction between immortal early embryonic cells and mortal differentiating derivative cells. Our approach is to optimize the use of systematic genomic approaches for profiling gene expression patterns by large-scale cDNA sequencing and cDNA microarray technology with a current cohort of ~22,000 genes collected from early mouse embryos and stem cells. We are particularly focusing on preimplantation development - the process of fertilized egg gradually losing its totipotency. In our previous work, we identified many genes that show stage-specific expression patterns during preimplantation mouse development. However, these genes have been identified by EST frequency, which is a relatively inaccurate and far from ideal way to do gene expression profiling. The cDNA microarray-based gene expression profiling will provide more reliable information. During this period, we have completed the global expression profiling of all preimplantation stages in mouse, which revealed and characterized the distinctive patterns of maternal RNA degradation and zygotic gene activation, including two major transient waves of de novo transcription. The first wave corresponds to zygotic genome activation (ZGA); the second wave, named mid-preimplantation gene activation (MGA), precedes the dynamic morphological and functional changes from the morula to blastocyst stage. Further expression profiling of embryos treated with inhibitors of transcription, translation, and DNA replication revealed that the translation of maternal RNAs is required for the initiation of ZGA. We propose a cascade of gene activation from maternal RNA/protein sets to ZGA gene sets and thence to MGA gene sets. The large number of genes identified as involved in each phase is a first step toward analysis of the complex gene regulatory networks. To start to examine whether these genes identified here might be involved in the immortal/mortal transition process, we are currently analyzing the nature of these genes in greater detail. We have also compared the global gene expression profiling between oocytes derived from young mouse and those derived from old mouse. This has characterized and identified the age-associated changes of oocytes at molecular levels. We are currently analyzing genes that we have identified in this study. Finally, we are also developing a high throughput whole-mount in situ hybridization technique on preimplantation embryos.
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