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Studies on developmental aberration of somatic clones by global gene expression analysis

通过全局基因表达分析研究体细胞克隆的发育畸变

基本信息

批准号：
16380192
负责人：
KONO Tomohiro
金额：
$ 10.05万
依托单位：
Tokyo University of Agriculture
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

项目摘要

Somatic cloning does not always result in ontogeny in mammals, and development is often associated with various abnormalities and embryo loss with a high frequency. This is considered to be due to aberrant gene expression resulting from epigenetic reprogramming errors. In the present study we conducted to explore the genes that are involved in developmental defects of somatic cloned mouse embryos. In order to address this issue, we carried out global screening of candidate genes by subtraction analysis using ES-cloned blastocysts, and differential display analysis using 2-cell embryos. From subtraction analysis 218 genes were detected as differentially expressed genes at blastocysts. Of these 198 clones were successfully sequenced and by BLAST search analysis 158 genes are detected as known genes. To confirm differentially expression, we carried out quantitative gene expression analysis by real-time PCR. The results showed that 5 of 10 genes were actually highly expressed in ES-cloned blastocysts. Further, we compared gene expression patterns using differential display RT-PCR (DDRT-PCR) between the NT and IVF embryos at the 2-cell stage to detect some abnormalities affecting later development of NT embryos. Aberrant gene expression was detected in NT embryos compared with IVF embryos, and MuERV-L and Dnaja2 genes were down-regulated and Inpp5b and Chst12 genes were up-regulated in the NT embryos. Further analysis showed that the expression of zygotically activated genes such as Interferon-?, Dub-1, Spz1, DD2106 and DD2111 were not properly activated in NT embryos, suggesting that the cellular process involved in the control of the zygotic genome activation is not appropriately regulated. These results indicate that abnormal gene expression has already occurred at the early stage of preimplantation development as a failure of nuclear reprogramming.

体细胞克隆并不总是导致哺乳动物的个体发育，发育通常与各种异常和胚胎丢失有关。这被认为是由于表观遗传重编程错误导致的异常基因表达。本研究旨在探索与体细胞克隆小鼠胚胎发育缺陷相关的基因。为了解决这一问题，我们进行了全面的候选基因的筛选，使用ES克隆囊胚差减分析，并使用2-细胞胚胎的差异显示分析。差减分析共检测到218个差异表达基因。其中198个克隆成功测序，通过BLAST搜索分析，158个基因被检测为已知基因。为了确认差异表达，我们通过实时PCR进行了定量基因表达分析。结果表明，10个基因中有5个在ES克隆囊胚中高表达。此外，我们使用差异显示RT-PCR（DDRT-PCR）的NT和IVF胚胎在2-细胞期之间的基因表达模式进行比较，以检测一些影响NT胚胎后期发育的异常。与IVF胚胎相比，NT胚胎中MuERV-L和Dnaja 2基因表达下调，Inpp 5 b和Chst 12基因表达上调。进一步的分析表明合子激活基因如干扰素-β，Dub-1，Spz 1，DD 2106和DD 2111在NT胚胎中没有被正确激活，这表明参与合子基因组激活控制的细胞过程没有被适当调节。这些结果表明，异常的基因表达已经发生在早期阶段的植入前的发展作为一个失败的核重编程。

项目成果

期刊论文数量（19）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Nuclei of oocytes derived from mouse parthenogenetic embryos are competent to support development to term.

来自小鼠孤雌胚胎的卵母细胞核能够支持足月发育。

DOI：
发表时间：
2004
期刊：
Bio Reprod. 71
影响因子：
0
作者：
Sato A;Kono T;Nakada K;Ishikawa K;Inoue S;Yonekawa H;Hayashi J.;Niwa K et al.
通讯作者：
Niwa K et al.

Current status of chromosomal abnormalities in mouse embryonic stem cell lines used in Japan.

日本使用的小鼠胚胎干细胞系染色体异常的现状。

DOI：
发表时间：
2006
期刊：
Comp Med. 56
影响因子：
0
作者：
Sugawara;A. et al.
通讯作者：
A. et al.

Pluripotential competence of cells associated with Nanog activity

DOI：
10.1016/j.mod.2004.08.008
发表时间：
2005-01-01
期刊：
MECHANISMS OF DEVELOPMENT
影响因子：
2.6
作者：
Hatano, S;Tada, M;Tada, T
通讯作者：
Tada, T

Gene therapy for progeny of mito-mice carrying pathogenic mtDNA by nuclear transplantation

DOI：
10.1073/pnas.0506197102
发表时间：
2005-11-15
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
影响因子：
11.1
作者：
Sato, A;Kono, T;Hayashi, JI
通讯作者：
Hayashi, JI

Abnormal gene expression in nuclear transfer embryos at the time of zygotic gene activation in mice.

小鼠合子基因激活时核移植胚胎中基因表达异常。

DOI：
发表时间：
2006
期刊：
MRD (in press)
影响因子：
0
作者：
Nakagawa;Y.;Yamada;N.;Shimizu;H.;Shiota;M.;Tamura;M.;Kim-Mitsuyama;S.;Miyazaki;H.;Wu Q et al.;Sugawara A et al.;Hiura H et al.;Ono Y et al.;Suzuki T et al.
通讯作者：
Suzuki T et al.