Demecolcine- and nocodazole-induced enucleation in mouse and goat oocytes for the preparation of recipient cytoplasts in somatic cell nuclear transfer procedures
Demecolcine- and nocodazole-induced enucleation in mouse and goat oocytes for the preparation of recipient cytoplasts in somatic cell nuclear transfer procedures
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DOI：
10.1016/j.theriogenology.2010.09.022
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发表时间：
2011-02-01
期刊：
影响因子：
2.8
通讯作者：
Ibanez, Elena
中科院分区：
文献类型：
--
作者：
Costa-Borges, Nuno;Teresa Paramio, Maria;Ibanez, Elena
用去甲氧胆碱(DEM)处理预激活的卵母细胞，可以诱导第二极体(PB2)内的所有卵母细胞染色体被挤出。然而，体细胞核移植(SCNT)后诱导去核(IE)率普遍较低，这些细胞质支持胚胎发育的能力受到损害。在这里，我们探索了用DEM或另一种抗有丝分裂药物诺康唑(NOC)进行短期处理是否能提高LE效率，并确定了在产生的细胞质中进行核移植的最合适时机。我们首次证明了用NOC可以在小鼠和山羊卵母细胞中完成IE，并且用DEM或NOC短期处理可以得到相似的IE率，这被证明是品系和物种特异性的。由于两种抗有丝分裂药物诱导的去核都是可逆的，IE方案与机械抽吸PB2相结合以提高小鼠卵母细胞的永久性去核率。由此产生的细胞质产生的克隆小鼠胚胎都没有发育到囊胚期。然而，当它们在激活和抗有丝分裂处理之前被重建时，它们的体外胚胎发育与机械去核卵母细胞产生的克隆胚胎相似。(C)2011 Elsevier Inc.保留所有权利。
Treatment of pre-activated oocytes with demecolcine (DEM) has been shown to induce the extrusion of all oocyte chromosomes within the second polar body (PB2). However, induced enucleation (IE) rates are generally low and the competence of these cytoplasts to support embryonic development following somatic cell nuclear transfer (SCNT) is impaired. Here, we explored whether short treatments with DEM or another antimitotic, nocodazole (NOC), improve LE efficiency, and determined the most appropriate timing for nuclear transfer in the cytoplasts produced. We show, for the first time, that IE can be accomplished in mouse and goat oocytes using NOC and that short treatments with DEM or NOC result in similar IE rates, which proved to be strain- and species-specific. Because enucleation induced by both antimitotic drugs is reversible, the IE protocol was combined with the mechanical aspiration of PB2s to increase permanent enucleation rates in mouse oocytes. None of the cloned mouse embryos produced from the resultant cytoplasts developed to the blastocyst stage. However, when they were reconstructed prior to the activation and antimitotic treatment, their in vitro embryonic development was similar to that of cloned embryos produced from mechanically-enucleated oocytes. (C) 2011 Elsevier Inc. All rights reserved.