Transfection of mammalian tissue culture cells
FuGENE 6 (Roche)
- Warm to room temperature before use
- Mix before use
- Do not aliquot
- Always pipet into serum-free medium, not into empty tubes
- Initial testing
- In certain cells antibiotics or serum present in the medium may have adverse effects
- Use FuGENE (µl):DNA (µg) amounts of 3:2, 3:1 and 6:1
Preparation of complex
- Pipette serum free medium into a tube, followed by the FuGENE6. Tap to mix.
- Add the DNA.
- Tap the tube to mix. DO NOT VORTEX.
- Dropwise, add the complex to the cells, distributing around the well. Swirl to ensure even dispersal.
- Return the cells to the incubator.
Table 1. Recommended amounts of components.
|Type of dish or plate||Surface area (cm2)||Total media volume (ml)||Dilution volume (µl)||Starting volume of FuGENE 6 (µl)||Starting mass of DNA (µg)|