Difference between revisions of "Lipofectamine Plasmid Transfection"
From Bridges Lab Protocols
Davebridges (Talk | contribs) m (→Protocol (6 well dish)) |
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==Protocol (6 well dish)== | ==Protocol (6 well dish)== | ||
| − | #Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics. | + | #Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics (DMEM with 10% FBS, no PSG). |
#For each well prepare: | #For each well prepare: | ||
##250uL OptiMEM plus 4 ug of DNA. | ##250uL OptiMEM plus 4 ug of DNA. | ||
Revision as of 14:45, 13 August 2009
Materials
- Cells in a 6 well dish, plated and at >90% confluence
- Lipofectamine 2000 (Invitrogen cat# 11668-019 for 1.5mL)
Protocol (6 well dish)
- Plate cells the day before so that they are at 90-95% confluence in 2 mL media without antibiotics (DMEM with 10% FBS, no PSG).
- For each well prepare:
- 250uL OptiMEM plus 4 ug of DNA.
- 250uL OptiMEM plus 10 uL of Lipofectamine.
- Incubate ~5 minutes at room temperature.
- Combine the two volumes of OptiMEM/DNA and OptiMEM/Lipofectamine.
- Incubate ~20 min at room temperature.
- Add DNA/Lipofectamine complexes (~500 uL) to the 2 mL of media on the cells
- Gently rock plate to mix
- After ~4h re-feed cells with normal media.
Protocol adapted from Product Manual
