Cloning of the transgenic pigs expressing human decay accelerating factor and N-acetylglucosaminyltransferase III
Cloning of the transgenic pigs expressing human decay accelerating factor and N-acetylglucosaminyltransferase III
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DOI：
10.1089/clo.2004.6.294
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发表时间：
2004-09-01
期刊：
影响因子：
--
通讯作者：
Nagashima, H
中科院分区：
文献类型：
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作者：
Fujimura, T;Kurome, M;Nagashima, H
本文介绍了利用重组糖链生物合成的人衰变加速因子(DAF，CD55)和N-乙酰氨基葡萄糖转移酶III(GnT-III)的转基因猪成纤维细胞生产克隆猪。采用两种核移植方案：两步激活法(TA)和延迟激活法(DA)。将去核的体外成熟卵母细胞和供体细胞分别用TA法和DA法在含钙或无钙的培养液中电融合，1~1.5h后进行电激活。TA法和DA法构建的核移植胚胎体外囊胚形成率分别为8%和14%。将每种方法获得的重构胚胎移植到受体猪体内，都产生了克隆仔猪。这些克隆猪表达转基因的数量与它们的核供体细胞一样多。综上所述，(1)猪的克隆可以通过TA或DA核移植的方法进行；(2)转基因动物的核供体细胞在克隆猪中可以保持转基因的表达。
The present paper describes production of cloned pigs from fibroblast cells of transgenic pigs expressing human decay accelerating factor (DAF, CD55) and N-acetylglucosaminyltransferase III (GnT-III) that remodels sugar-chain biosynthesis. Two nuclear transfer protocols were used: a two-step activation (TA) method and a delayed activation (DA) method. Enucleated in vitro-matured oocytes and donor cells were electrically fused in a calcium-containing medium by TA method or in a calcium-free medium by DA method, followed by electrical activation 1-1.5 h later, respectively. In vitro blastocyst formation rates of nuclear transferred embryos reconstructed by TA and DA method were 8% and 14%, respectively. As a result of embryo transfer of the reconstructed embryos made by each method into recipient pigs, both gave rise to cloned piglets. These cloned pigs expressed transgene as much as their nuclear donor cells. In conclusions, (1) pig cloning can be carried out by TA or DA nuclear transfer methods, (2) expression of transgenes can be maintained to cloned pigs from the nuclear donor cells derived from transgenic animals.