Cell transfection is the introduction of nucleic acids such as DNA and RNA into eukaryotic cells through non-viral techniques to create genetically modified cells. By modifying the genetic content of cells in transfection medium, you can efficiently study cellular processes and gene function.
Some factors that affect transfection efficiency
- Cell health. To optimize your culture processes, be sure to grow cells in appropriate culture media for your specific cell line. Do your cells have contaminants like microorganisms or mycoplasma? Then discard them and reseed from uncontaminated stock. Also, supplement transfection medium with growth factors or other additives as needed to enhance viability. If you use serum be sure to consider how that may affect transfection efficiency.
- Confluency. Another important factor that influences transfection is confluency. This refers to the coverage of the culture growth surface, such as a tissue culture flask, by cells. Cells should be actively dividing and healthy for best results. A good target is 50% to 80% confluency.
- DNA quantity and quality. The optimal amount of DNA for efficient transfection varies based on the DNA type, transfection medium, number of cells, and cell type, among other factors. Plasmid DNA used in transfection processes should contain minimal RNA, protein, and contaminants.
How to improve cell transfection efficiency
Transfection efficiency largely depends on the type of cell and conditions in the cell culture vessel. Here are some techniques used to enhance efficiency:
- Optimize the lipid to DNA ratio if using a lipid as transfection reagent: Ensure the positive charge from the cationic lipid component in transfection medium equals or exceeds the negative charge from the DNA.
- Modulate the amount of DNA: Perform a study to determine the recommended DNA and reagent concentration ranges for peak efficiency.
- Keep track of transfection time: Evaluate different transfection times to find the one that gives the best efficiency for your cell lines.
- Consider whether you can remove serum: Since serum may interfere with the transfection, opt for serum-free transfection medium to obtain peak results.
Below are answers to some frequently asked questions regarding transfection.
What are the two types of transfection?
Based on the long-term effect on cells, transfection techniques come in two broad classes, transient and stable. In transient transfection, foreign DNA doesn’t integrate with chromosomes from the host cells. On the other hand, stable transfection integrates the DNA of interest into the host genome. As a result, descendants of these cells also express the new gene.
What is transfection used for?
Industrial uses of transfection include the delivery of DNA or RNA sequences into cells for biomanufacturing. This process also introduces genes into viral vectors, such as lentivirus and adeno-associated virus (AAV), for recombinant vaccine production and for use in cell therapy and gene therapy.