I am at the point where I have to transfect my cells and I read about both electroporation and chemical transfections. I have a fairly large plasmid (20Kb) and I wanted your opinion about the best way to transfect my cells.

Answer

Great question to start this “ask the expert”. Yes both electroporation and chemical transfection are the two most popular methods for moving DNA into cells. The advantage of electroporation is that you can get DNA into all cell types and it is especially good for eukaryotic cells and stem cells. It works by using a very short pulse of high-voltage passed through a cuvette containing cells in a special conductive medium. It is important to optimize to minimize cell death, but with optimization done, you might get close to 100% efficiency with certain cell types,but not all. Of course you need an electroporation device of some kind. In case you need one Neon by Life Technologies is a good choice since it is simple to use and set up. Chemical transfection using cationic lipids is probably what people use most at this time for transfection. It does not require special equipment. One of the possible drawbacks is thatt you can’t transfect all cell types. Another potential problem is that there is not one cationic lipid that works on all cell types, so you do have to do some digging to learn what specific cationic lipid will work for your cell type. My suggestion is that you should go to something like the Life Technologies transfection collection on their website, there you will find recommendations for many different cell types and references. I will mention that regardless of which method you choose, follow the protocols and do not skip any steps, equilibrations of reagents into transfection medium is important. Also, remember that optimization is important for your cell type. The same cell type grown in different labs might transfect with a different efficiencies because of the history of the cells.