Simple, Efficient, and Reproducible Gene Transfection of Mouse Embryonic Stem Cells by Magnetofection

Embryonic stem (ES) cells are recognized as an excellent cell culture model for studying develop-mental 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 af-fecting 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 abil-ity to form embryoid bodies and differentiated in vitro into cells of the three germ layers. eGFP ex-pression was sustained during stem cell proliferation and differentiation. This is the first transfec-tion 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. 13