| 绵羊羊水干细胞外源基因高效转染及快速筛选方法的建立 |
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| 引用本文： | 王凤武,郝斐,魏著英,刘东军,荣威恒. 绵羊羊水干细胞外源基因高效转染及快速筛选方法的建立[J]. 中国畜牧兽医, 2013, 40(11): 16-20 |
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| 作者姓名： | 王凤武 郝斐 魏著英 刘东军 荣威恒 |
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| 作者单位： | 1. 内蒙古农业大学动物科学学院, 内蒙古呼和浩特 010018;2. 内蒙古农牧业科学院, 内蒙古呼和浩特 010031;3. 内蒙古大学国家动物转基因技术中心, 内蒙古呼和浩特 010021 |
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| 基金项目： | 内蒙古自然科学基金(2011MS0410)。 |
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| 摘 要： | 试验旨在建立绵羊羊水来源多潜能干细胞(amniotic fluid-derived stem cells,AFSC)外源基因高效转染及快速筛选方法,同时保持其多潜能特性。采用脂质体法,以pCMV-DsRed为报告质粒,转染绵羊AFSC,36 h后,流式细胞仪分析外源基因的瞬时转染效率为69.17%。以浓度为6 mg/mL的G418筛选5 h获得纯化转基因绵羊AFSC单克隆。转基因绵羊AFSC RT-PCR检测表达Oct-4、SSEA-1,体外悬浮培养可形成类胚体。结果表明,转基因标记绵羊AFSC保持了发育全能性的潜能,构建的转基因绵羊AFSC可以示踪性研究其在体内的分化路径和最终宿主。
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| 关 键 词： | 绵羊 羊水干细胞 报告质粒 基因转染 高效转染 快速筛选 |
| 收稿时间： | 2013-04-18 |
Exogenous Gene Efficient Transfection in vitro and Rapid Screening of Sheep Amniotic Fluid Stem Cells |
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| WANG Feng-wu,HAO Fei,WEI Zhu-ying,LIU Dong-jun,RONG Wei-heng. Exogenous Gene Efficient Transfection in vitro and Rapid Screening of Sheep Amniotic Fluid Stem Cells[J]. China Animal Husbandry & Veterinary Medicine, 2013, 40(11): 16-20 |
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| Authors: | WANG Feng-wu HAO Fei WEI Zhu-ying LIU Dong-jun RONG Wei-heng |
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| Affiliation: | 1. College of Animal Science, Inner Mongolia Agricultural University, Hohhot 010018, China;2. Inner Mongolia Academy of Agricultural and Animal Husbandry, Hohhot 010031, China;3. The National Animal Transgenic Technology Centre, Inner Mongolia University, Hohhot 010021, China |
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| Abstract: | The study was designed to establish a method for the efficient gene transfection in vitro and rapid screening of amniotic fluid-derived stem cells (AFSC), while maintaining their pluripotency. pCMV-DsRed was used as the reporter plasmid to transfect amniotic fluid-derived stem cells by liposome. 36 h later, transient transfection efficiency was 69.17% by flow cytometry. The purified transgenic sheep AFSC monoclones were obtained through the selection of G418 at the concentration of 6 mg/mL after the treatment of 5 h. The transgenic sheep AFSC was positive for Oct-4, SSEA-1 by RT-PCR. AFSC could also formed embryoid bodies by in vitro suspension culture. The results implied a possibility that the transgenic sheep AFSC could be used to track down in vivo differentiation path and the definitive host. |
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| Keywords: | sheep amniotic fluid stem cells reporter plasmid gene transfection efficient transfection rapid screening |
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