Observing and Manipulating Pluripotency in Normal and Cloned Mouse Embryos

The mouse ooplasm is the ideal platform to study and compare induced and natural pluripotency because it can support both, after somatic cell nuclear transfer (cloning) and after fertilization, respectively. The amount of pluripotency induced after cloning is variable but always limited compared to fertilization. It can be visualized conveniently if the nucleus donor cells carry a green fluorescent protein (GFP) reporter under control of the pluripotency-associated gene Oct4 promoter. Thus we produced cloned and fertilized mouse embryos transgenic for Oct4-GFP (GOF18-∆PE-EGFP). We also developed and validated a live cell imaging method, whereby we resolve and selectively pick cloned embryos that hold distinct amounts of induced pluripotency as predicted by GFP intensity and measured by embryonic stem cell derivation. Currently we are developing a microinjection method to change the level of Oct4 without modifying the genome of the embryo. Here we discuss our findings in relation to the epigenetic reprogramming of the nucleus transplant and to cell fate decisions in the cloned or fertilized mouse embryo.