Simple, efficient, and reproducible gene transfection of mouse embryonic stem cells by magnetofection
Simple, efficient, and reproducible gene transfection of mouse embryonic stem cells by magnetofection
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DOI：
10.1089/scd.2007.0064
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发表时间：
2008-02-01
影响因子：
4
通讯作者：
Park, Se Pill
中科院分区：
文献类型：
--
作者：
Hyun, Chang;Young, Eun;Park, Se Pill
胚胎干细胞是研究胚胎发育机制和治疗调控的理想细胞培养模型。这项工作的目的是确定是否使用磁转染是一种有效的方法来操纵干细胞的遗传，而不会对它们的增殖或自我更新能力产生不利影响。我们将我们的磁转染结果与使用FuGENE 6的保守方法的结果进行了比较。使用增强型绿色荧光蛋白（eGFP）作为D3小鼠ES（mES）细胞中的报告基因，我们发现磁转染在干细胞中的基因递送效率（45%）显著高于FuGENE 6方法（15%），而两者在NIH-3 T3细胞中均显示出有效的转染（60%）。尽管转染的D3（D3-eGFP）mES细胞经历了大量传代（> 50），但高百分比的细胞保留ES标记物，例如Oct-4和阶段特异性胚胎抗原-1（SSEA-1）。它们还保留了形成胚状体的能力，并在体外分化成三个胚层的细胞。eGFP表达在干细胞增殖和分化期间持续。这是第一个在ES细胞中使用磁转染的转染报告。根据我们的结果，我们得出结论，磁转染是一种有效的和可靠的外源DNA导入小鼠ES细胞的方法，并可能成为首选的方法。
Embryonic stem (ES) cells are recognized as an excellent cell culture model for studying developmental mechanisms and their therapeutic modulations. The aim of this work was to define whether using magnetofection was an efficient way to manipulate stem cells genetically without adversely affecting their proliferation or self-renewal capacity. We compared our magnetofection results to those of a conservative method using FuGENE 6. Using enhanced green fluorescent protein (eGFP) as a reporter gene in D3 mouse ES (mES) cells, we found that magnetofection gave a significantly higher efficiency (45%) of gene delivery in stem cells than did the FuGENE 6 method (15%), whereas both demonstrated efficienct transfection in NIH-3T3 cells (60%). Although the transfected D3 (D3-eGFP) mES cells had undergone a large number of passages (> 50), a high percentage of cells retained ES markers such as Oct-4 and stage-specific embryonic antigen-1 (SSEA-1). They also retained the ability to form embryoid bodies and differentiated in vitro into cells of the three germ layers. eGFP expression was sustained during stem cell proliferation and differentiation. This is the first transfection report using magnetofection in ES cells. On the basis of our results, we conclude that magnetofection is an efficient and reliable method for the introduction of foreign DNA into mouse ES cells and may become the method of choice.