Abstract

MORO, L.N.; VICHERA, G.; OLIVERA, R.  and  SALAMONE, D.. Evaluation of demecolcine assisted enucleation as a method to avoid the UV exposure in the bovine embryos production by cloning. InVet [online]. 2010, vol.12, n.2, pp.195-204. ISSN 1668-3498.

Cloning is a powerful scientific and economical tool, though inefficient. The ultraviolet irradiation (UV) used for oocytes enucleation generates structural and functional changes, affecting the viability of reconstructed embryos. The microtubule inhibitor demecolcina (DMC) permits the production of a cytoplasmic protrusion containing the oocyte nucleus, avoiding the UV irradiation for its identification and removal. Initially, oocytes were exposed to 0,4μg/ml DMC during whole in vitro maturation (24h) or after 21h, obtaining 61,4% and 87% of the nuclei inside a visible protrusion, respectively. In a second experiment oocytes were incubated in 0,4μg/ml DMC after ionomycin exposure, with 52,7% efficiency. Finally, protrusion nuclei formed after the best treatment (DMC21h) were removed, without UV radiation. Each enucleated and zona pelucida free oocyte was electro-fused with a blastomere from a 8 cell IVF embryo. Control group was not exposed to DMC and UV was used for the enucleation procedure. No statistical differences were observed in cleavage (67% vs 76%) nor in blastocyst rates (7,4% vs 3,8%) between both groups, although enucleation was technically easier using the new treatment

Keywords : Cloning; Demecolcine; Ultraviolet radiation; Protrusion.

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