stable transfection of mouse embryonic stem cells - help! (Dec/21/2005 )
i am trying to transfect mouse embryonic stem cells by electroporation of a linearized plasmid, in which expression is driven by the CMV promoter. following G418 selection, i was unable to detect any expression of the protein of interest by FACS. has anyone tried other transfection methods (e.g. lipofectamine, neophectin) with better success? any suggestions would be greatly appreciated!!
Before going to create a stable transformation, why don't you try to transfect a reporter plasmid (for instance with GFP as you have can FACS) today en try and detect reporter gene expression tomorrow or the day after?
Have you determined the concentration of G418 that kills non-transfected cells?