The TransMessenger Transfection Reagent from Qiagen is designed for the efficient transfection of eukaryotic cells with RNA (including siRNA). The reagent works using principles similar to the liposome-based DNA transfection reagents that Qiagen also makes. In this case, the RNA is “condensed” using an “Enhancer” that is then mixed with the TransMessenger reagent, which forms a complex with the condensed RNA. The complex of RNA and TransMessenger is then mixed with serum-free media and incubated with target cells. The efficiency of the transfection depends on a number of factors including cell density, the amount of RNA used and the amount of time the cells are exposed to the TransMessenger-RNA complexes. The protocol for siRNA transfection does not differ significantly from the standard RNA transfection procedure.
We purchased the TransMessenger reagent for the high-throughput transfection of siRNAs for gene silencing studies using a transfected-cell array. Our choice was initially dictated by our previous good experiences with Qiagen products for plasmid purification and RNA isolation. We expected extensive troubleshooting using the reagent the first time, however, to our surprise, we achieved a gene silencing effect during the first experiments. One of the factors that we have come to realize as being extremely important is that a proper ratio of siRNA to TransMessenger reagent is critical for successful transfections. Another factor that strongly influences transfection efficiency is the cell density at transfection, which should not be too high. Once optimized, we recommend keeping the same conditions for a particular cell type for future experiments, that is, the same number of cells should be seeded for each experiment. We have also found that the RNA/TransMessenger ratio as well as the cell’s optimal exposure time are both strongly cell-type specific. In our hands, the conditions with which we achieved excellent transfection rates for HEK cells could not be used for HeLa cells. One nice surprise was that although the reagent is designed for the transfection of RNA in suspension, we successfully used TransMessenger for transfecting siRNAs spotted on a glass surface (although we modified the protocol slightly).
Gene-silencing studies based on the recently described RNA interference (RNAi) phenomenon are currently expanding at a rapid pace and this field has become very competitive for researches involved in it. On the other hand, siRNA chemistry and biology is still not well characterized. The TransMessenger Reagent, through its reliability and simplicity of use, provides a shortcut for those who are more interested in the physiological effects of their specific RNAi rather than in the siRNA mechanism itself.
Michal Janitz, M.D.
Max Planck Institute for Molecular Genetics
Berlin, Germany