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標題: Transgenic cloned mice expressing enhanced green fluorescent protein generated by activation stimuli combined with 6-dimethylaminopurine
作者: Chen, C.H.
Stone, L.
Ju, J.C.
Lien, W.T.
Liu, M.S.
Tu, C.F.
Lee, K.H.
關鍵字: parthenogenetic mouse embryos;repetitive calcium transients;somatic-cell nuclei;bovine oocytes;map kinase;stem-cells;cloning;culture;inhibition;egg
Project: Reproduction in Domestic Animals
期刊/報告no:: Reproduction in Domestic Animals, Volume 43, Issue 5, Page(s) 547-555.
Most studies of mouse cloning successfully achieved activation of the reconstructed oocytes by strontium (Sr) combined with cytochalasin B (CB) treatment. A protein kinase inhibitor, 6-dimethylaminopurine (6-DMAP), was used to inhibit the activity of maturation promoting factor for activation of oocytes, but it has never been successfully applied in mouse cloning. This study investigates the activation efficiency of 6-DMAP in mouse somatic cell nuclear transfer (SCNT). Higher parthenogenetic blastocyst rates (71-72%, p < 0.05) were achieved in the oocytes treated with Sr6D (10 mM Sr combined with 2 mM 6-DMAP for 4 h) and Sr6D + SrCB (Sr6D for 2 h then Sr combined with 5 mu g/ml CB for another 2 h), and a higher rate of hatching and hatched blastocyst was observed in the Sr6D + SrCB group (31%, p < 0.01) compared with other treatment groups (1-8%). For mouse cloning, cumulus cells of enhanced green fluorescent protein (EGFP)-expressed ESC chimera F1 were used as donor nuclei. Following activation, better development of the cloned embryos was observed in Sr6D + SrCB treatment. Moreover, different media, i.e. KSOM-AA, MEM-alpha and MK, for culturing cloned embryos were also compared in this study. Better morula/blastocyst (40%) and blastocyst (29%) rates were achieved in the embryos cultured in MEM-alpha medium (p < 0.05). Consequently, four EGFP cloned mice were generated in the activation treatment containing 6-DMAP following embryo transfer. In conclusion, treatment with 6-DMAP in combination with other activation stimuli successfully activates mouse reconstructed oocytes and support full-term development of the transgenic SCNT cloned embryos.
ISSN: 0936-6768
DOI: 10.1111/j.1439-0531.2007.00951.x
Appears in Collections:動物科學系

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