- Old cells transfect poorly - optimal transfection occurs when passage number is < 30.
- Lipofectamine reagent is less effective (and can eventually go bad) if allowed to rise about 4C too often. Leave Lipofectamine in the fridge until you are ready to add it to the tubes, and replace it immediately after use.
- After adding the DNA to the Lipofectamine (LTX), incubate for 20 minutes (original protocol calls for 5 minutes).
- Make 100uL aliquots of midipreps (to reduce the number of freeze thaw cycles and prevent loss of DNA supercoiling).
- Use tagBFP (instead of eBFP) as transfection marker.
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