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Generation of cloned mice and application on the field of laboratory animal science

克隆小鼠的产生及其在实验动物科学领域的应用

基本信息

批准号：
11308034
负责人：
ITO Mamoru
金额：
$ 25.29万
依托单位：
Central Institute for Experimental Animals
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11308034/
关键词：
experimental animals cloned mice nuclear transfer embryonic stem cells mitochondria ヘテロプラスミー

项目摘要

In order to study the practical application of cloning technology to the field of laboratory animal science, we have attempted to generate cloned mice from fibroblasts and embryonic stem cells by a serial nuclear transfer technique, and also examined the characteristics of the cloned mice. We successfully generated 5 ($ % of blastocysts transferred) mice from metaphase-arrested CD1 or (CD1 x B6) F1 fetal fibroblasts, and 27 (6%) and 20 (2%) mice from two TT2 ES cell lines that were independently inactivated different genes, G9a and Oviduct-specific glycoprotein (OGP). In these cloned mice, the commonly observed morphological abnormality was hypertrophy of the placenta, which was over 2-5 times over than the controls. In addition, some cloned mice showed increased body weights and open eyelids at birth. However, these abnormalities have not transmitted the progeny by mating, indicating that the abnormalities seen in the cloned mice were caused by inappropriate reprogramming of the epigenetic modifications, but not by accumulative DMA mutations, and was correctly reprogrammed through the germ line. In fact, we clarified that the expression pattern of imprinting genes, i.e., Igf2 and H19, were distorted in the cloned mice and their placentas. However, the fact that the progeny from the cloned mice that showed various abnormalities were normal suggests the usefulness of cloning technology in the field of laboratory animal science. The possibilities in which transfer of mitochondria originated from donor cells may influence the phenotype of the clone mice were also demonstrated.

为了研究克隆技术在实验动物科学领域的实际应用，我们尝试了用连续核移植技术从成纤维细胞和胚胎干细胞中产生克隆小鼠，并检查了克隆小鼠的特征。我们成功地从中期阻滞的CD 1或（CD 1 x B6）F1胎儿成纤维细胞中产生了5只（囊胚转移的$ %）小鼠，从两种TT 2 ES细胞系中产生了27只（6%）和20只（2%）小鼠，这两种细胞系分别是不同基因、G9 a和输卵管特异性糖蛋白（OGP）失活的。在这些克隆小鼠中，常见的形态异常是胎盘肥大，是对照组的2-5倍以上。此外，一些克隆小鼠在出生时体重增加，眼睑张开。然而，这些异常并没有通过交配传递给后代，这表明在克隆小鼠中观察到的异常是由表观遗传修饰的不适当重编程引起的，而不是由累积的DMA突变引起的，并且通过生殖系正确地重编程。事实上，我们阐明了印记基因的表达模式，即，Igf 2和H19在克隆小鼠及其胎盘中发生畸变。然而，克隆小鼠的后代表现出各种异常却正常，这一事实表明克隆技术在实验动物科学领域是有用的。还证明了来自供体细胞的线粒体的转移可能影响克隆小鼠的表型。