Cell transfection
100 cm dishes
Transfection reagents for PA-1:
X-tremeGENE™ 9 DNA Transfection Reagent
Opti-MEM
The day before transfection, trypsinize the cells, count and seed them.
Incubate at 37ºC for at least 16-18 h prior to transfection (no longer than 20 h).
For a 100 cm dish:
- In a sterile tube, add 485 ul of room temperature (RT) warmed Opti-MEM.
- Add 15 µl of X-tremeGENE™ 9 to the tube (drop by drop, directly over Opti-MEM) and incubate at RT for 5 minutes (up to 20 min).
- In another sterile tube, add 5 µg of plasmidic DNA.
- Once incubation is finished, add the Opti-MEM/ X-tremeGENE™ 9 mix to the tube with the DNA.
Flick the tube to ensure everything is mixed and incubate at RT for 20 min (up to 60 min).
- Drop by drop, add the mix to the cells.
- Incubate overnight at 37 ºC.
- Change the culture media the next day.