Promoter, transgene, and cell line effects in the transfection of mammalian cells using PDMAEMA-based nano-stars

• 4 cell lines, 4 promoters, and 3 gene products were studied, i.e. 48 combinations.
• Distinct cell line depended effects were observed.
• Jurkat cell results tended to differ from that obtained with the other cells.
• Co-transfection works well in CHO cells, but fails in up to 80% of Jurkat cells.
• High transfection efficiency in CHO and HEK cells is maintained in spite of pDNA dilution.

Non-viral transfection protocols are typically optimized using standard cells and reporter proteins, potentially underestimating cellular or transgene effects. Here such effects were studied for two human (Jurkat, HEK-293) and two rodent (CHO-K1, L929) cell lines and three fluorescent reporter proteins. Expression of the enhanced green fluorescent protein (EGFP) was studied under the control of the human elongation factor 1 alpha promoter and three viral promoters (SV40, SV40/enhancer, CMV), that of ZsYellow1 (yellow fluorescence) and mCherry (red fluorescence) for the CMV promoter. Results varied with the cell line, in particular for the Jurkat cells. Pair-wise co-transfection of the CMV controlled transgenes resulted in a significant fraction of monochromatic cells (EGFP for EGFP/YFP and EGFP/RFP co-transfections, YFP in case of YFP/RFP co-transfections). Only Jurkat cells were almost incapable of expressing YFP. Dilution of the plasmid DNA with a non-expressed plasmid showed cell line dependent effects on transfection efficiency and/or expression levels.

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