ThermoFisher
Lipofectamine 3000
L3000-015
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Transient transfection of plasmids into mammalian cells is an essential step in any research lab. While many reagents are widely available, Lipofectamine 3000 has become one of the most commonly used reagents when it comes to transient transfection. Here, I describe our approach to using this reagent when transfecting HCT116 cells.
Transient Transfection
HCT116 cells
Protocol for transfecting one well of a 6-well plate that is ~80% confluent using 1 µg of plasmid DNA. Prepare Tube 1 and 2 separately. Tube 1: 1 µg DNA + 125 µL OptiMEM + 4 µl P3000 reagent (Yellow cap). Tube 2: 125 µl OptiMEM + 4 µl Lipo3000 reagent (Red cap). Wait 5 mins and then mix Tube 1 and 2 together. Keep for another 15 mins and then add drop-wise to your cells.
Try to set up mastermixes when transecting multiple plasmids.
Transfection efficiency was assessed by WB for a transgene 24 hrs post transfection. The transgene was robustly expressed.
N/A
Works well on many types of cells and fairly easy to use.
Expensive and requires setting up two individual tubes for the transfection
High quality transfection reagent that maximizes transfection efficiency.