Assisted reproductive technologies (ARTs) in domestic animals are used to increase the efficiency of cattle production, especially by reducing the generation interval in breeding programs. Somatic cell nuclear transfer (SCNT) allows for the generation of identical individuals of high genetic merit and it is also used to generate transgenic animals by modification of the donor's genome. However, SCNT efficiency is still very low when compared to other ARTs such as in vitro fertilization. The use of embryonic stem cells (ESCs) as donor nucleus could enhance the outcome of SCNT as well as allowing targeted gene modification. Nevertheless, true ESCs were not derived so far in the bovine species. Thus, we focused on SCNT as an ART that can accomplish not only the horizontal propagation of a given genome but also introduce genome modifications. During SCNT, a somatic cell must be transformed into a pluripotent cell first - in a matter of hours - and then reinitiate embryonic differentiation. The first differentiation event the pre-implantation embryo is lineage specification, in which a fraction of the cells in the embryo will form the trophectoderm and the rest will form the fetus itself. This process is poorly understood in the bovine, not only in the context of SCNT but in fertilized embryos as well. We suggest that studying the mechanisms of lineage specification in bovine could help understand some of the inefficiencies observed in bovine SCNT and it could also help us develop novel strategies to obtain true bovine ESC. We focused in three genes, OCT4, CDX2 and SOX2 that are key regulatory factors in most pre-implantation mammalian embryos. First we tested the hypothesis that expression of OCT4 in donor cells would improve the efficiency of SCNT. Subsequently we tested the hypothesis that CDX2 is not required for trophectoderm establishment, but important in maintaining its integrity in bovine embryos. To further comprehend lineage specification in bovine, we tested the hypothesis that SOX2 is required for inner cell mass formation. We found that indeed preconditioning the somatic cells with OCT4 expression prior to SCNT has a distinct effect on the phenotype of the cloned blastocysts. We also found, contrary to our expectations, that CDX2 and SOX2 are not required for the first cell differentiation event in the bovine fertilized embryo. Overall, the data presented here has direct implications in the understanding of bovine embryology. It could help improve SCNT outcome and further understand lineage specification in the pre-implantation embryo as well as subsequent embryonic processes.
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