Early development of cloned bovine embryos produced from oocytes enucleated by fluorescence metaphase II imaging using a conventional halogen-lamp microscope.
Early development of cloned bovine embryos produced from oocytes enucleated by fluorescence metaphase II imaging using a conventional halogen-lamp microscope.
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DOI：
10.1089/cell.2014.0086
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发表时间：
2015-03
影响因子：
1.6
通讯作者：
D. Iwamoto;K. Yamagata;M. Kishi;Yoko Hayashi-Takanaka;H. Kimura;T. Wakayama;K. Saeki
中科院分区：
文献类型：
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作者：
D. Iwamoto;K. Yamagata;M. Kishi;Yoko Hayashi-Takanaka;H. Kimura;T. Wakayama;K. Saeki
受体卵母细胞的去核是体细胞核移植的关键步骤之一。然而，特别是在牛种中，在卵质中扩散的脂滴妨碍中期II（MII）染色体的鉴定和去核，从而克隆的成功率仍然很低。在这项研究中，我们使用了一种新的实验系统，使荧光观察活卵母细胞中的染色体没有任何损害。我们成功地可视化和去除成熟牛卵母细胞中的MII染色体。该实验系统包括注射与染色体结合的荧光标记抗体缀合物和使用常规卤素灯显微镜进行荧光观察。随着抗体浓度的增加，牛体外受精（IVF）后胚胎的卵裂率和囊胚率降低（p0.05）。对于SCNT胚胎的生产，新系统可以用作不透明卵母细胞（如反刍动物）中中期板位置的可靠预测器。
Enucleation of a recipient oocyte is one of the key processes in the procedure of somatic cell nuclear transfer (SCNT). However, especially in bovine species, lipid droplets spreading in the ooplasm hamper identification and enucleation of metaphase II (MII) chromosomes, and thereby the success rate of the cloning remains low. In this study we used a new experimental system that enables fluorescent observation of chromosomes in living oocytes without any damage. We succeeded in visualizing and removing the MII chromosome in matured bovine oocytes. This experimental system consists of injecting fluorescence-labeled antibody conjugates that bind to chromosomes and fluorescent observation using a conventional halogen-lamp microscope. The cleavage rates and blastocyst rates of bovine embryos following in vitro fertilization (IVF) decreased as the concentration of the antibody increased (p0.05). For the production of SCNT embryos, the new system can be used as a reliable predictor of the location of metaphase plates in opaque oocytes, such as those in ruminant animals.