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Selection of donor nuclei in somatic cell-mediated gene transfer using a co-transfection method
Ma, Hairong; Lu, Zili; Sun, Yi; Peng, Tao; Shuai, Zhiqiang; Ma, Yufang; Zhang, Yan; Wang, Liang; Cao, Xu; Wang, Hanqing
2007
发表期刊JOURNAL OF REPRODUCTION AND DEVELOPMENT
ISSN0916-8818
卷号53期号:1页码:95-104
摘要In this study, we introduced a co-transfection method for the selection of donor nuclei in somatic cell-mediated nuclear transfer. Two vectors were constructed in our experiment. One was pMSCV-GFP carrying the neomycin-resistant gene (Neo(r)) and the green fluorescent protein (GFP) reporter gene; the other was pBC1-GFP carrying the mammary gland-specific promoter and target gene GFP. Ovine adult fibroblasts were co-transfected with pMSCV-GFP and pBC1-GFP. The data from this work demonstrated that the GFP genes in both vectors could successfully co-integrate into the genomes of ovine adult fibroblasts in three of the four transgenic cell clones assayed. Furthermore, PCR analysis of transgenic embryos proved that the GFP genes in both vectors could cointegrate into the genomes of the reconstructed embryos. Subsequently, analysis of the developmental rate of the reconstructed embryos after nuclear transfer indicated that the blastocyst rate from the co-transfected donor cells was similar (approximate 8 percent) to that from individual pMSCV-GFP transfected donor cells. The influence of co-transfection resulting in modification of donor nuclei on development of reconstructed embryos was also investigated. The results of flow cytometric analysis indicated that the co-transfected ovine fibroblasts had similar quiescent characteristics in terms of cell cycle (G0+G1 percent: 73.20 +/- 4.04) to the individual pMSCV-GFP transfected fibroblasts (G0+G1 percent: 70.77 +/- 1.19) after they were treated with serum starvation for five days. Our results suggest that the co-transfection method can be used for selection of donor cell clones in somatic cell-mediated gene transfer experiments. It can be potentially extended to applications related to expression of functional protein in mammary glands and other transgenic research relevant to nuclear transfer.
关键词Cell Cycle Co-transfection Mammary Gland-specific Promoter Ovine Adult Fibroblast Somatic Cell-mediated Nuclear Transfer
DOI10.1262/jrd.18082
收录类别SCI
WOS记录号WOS:000244465700011
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文献类型期刊论文
条目标识符http://ir.xjipc.cas.cn/handle/365002/4122
专题资源化学研究室
作者单位Chinese Acad Sci, Xinjiang Tech Inst Phys & Chem, Urumqi 830011, Peoples R China;Chinese Acad Sci, Lanzhou Inst Chem Phys, Lanzhou 730000, Peoples R China;Xinjiang Golden Cattle Biol Incorp, Urumqi 830011, Peoples R China;Xinjiang Med Univ, Urumqi 830011, Peoples R China;Univ Alabama, Dept Pathol, Birmingham, AL 35294 USA;China Agr Univ, State Key Labs Agrobiotechnol, Coll Biol Sci, Beijing 475001, Peoples R China
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Ma, Hairong,Lu, Zili,Sun, Yi,et al. Selection of donor nuclei in somatic cell-mediated gene transfer using a co-transfection method[J]. JOURNAL OF REPRODUCTION AND DEVELOPMENT,2007,53(1):95-104.
APA Ma, Hairong.,Lu, Zili.,Sun, Yi.,Peng, Tao.,Shuai, Zhiqiang.,...&Wang, Hanqing.(2007).Selection of donor nuclei in somatic cell-mediated gene transfer using a co-transfection method.JOURNAL OF REPRODUCTION AND DEVELOPMENT,53(1),95-104.
MLA Ma, Hairong,et al."Selection of donor nuclei in somatic cell-mediated gene transfer using a co-transfection method".JOURNAL OF REPRODUCTION AND DEVELOPMENT 53.1(2007):95-104.
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