Optimized cationic lipid‐based gene delivery reagents for use in developing vertebrate embryos

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Abstract
We have used cationic lipid-based transfection reagents for ectopic gene expression experiments in developing vertebrate embryos. Lipofectamine, Lipofectamine 2000, and Lipofectamine enhanced with a disulfide linked pegylated lipid (mPEG-SS-DOPE) were initially tested and optimized in cell culture. Two reagent formulations, 1:4 (DNA:Lipofectamine 2000) Lipofectamine 2000, and 7.5% pegylated Lipofectamine, produced the highest levels of gene expression in vitro. Those formulations, containing the enhance green fluorescent protein reporter gene, were microinjected into intact vertebrate embryos—systemically through the vasculature and locally into selected tissues—to assess in vivo transfection efficiency. Whereas both formulations are capable of transfecting cells in developing embryos in vivo, greater transfection efficiencies in a broader range of tissue types were obtained with the pegylated Lipofectamine formulation. We conclude that in developing vertebrate embryos, optimized cationic lipid-based reagents are capable of producing significant levels of ectopic gene expression and can be used as alternatives to electroporation and viral-mediated gene delivery. Developmental Dynamics 235:2210–2219, 2006.