Description
GenJet™ DNA In Vitro
Transfection Reagent for MEFs is pre-optimized and conditioned for transfecting
Mouse Embryonic Fibroblasts (MEFs).
MEFs are required to support the growth of undifferentiated mouse or
human ES cells, iPS cells, thus they are also called feeder cells. MEFs
are isolated from mouse embryos and are used at their early passages.
Before MEFs are used as feeder cells,
they must be treated by
irradiation or mitomycin C to stop the cells from further dividing.
However MEFs are nortoriously hard to transfect.
With the pre-optimized reagent, you will get up to 85% GFP+ MEFs.
The "Shaved Cell
Transfection" protocol was optimized to achieve maximal transfection efficiency. Refer to the following optimal transfection conditions for maximal transfection
efficiency on MEFs cells. GenJet™ reagent, 1.0 ml, is sufficient for 500 transfections in 24 well plates or
250 transfections in 6 well plates.
|
Summary of
Optimal Shaved CellTransfection Conditions:
Cell culture
medium
Confluency
of cell on the
day of shaving
Cell density for plating
GenJet™ (µl) : DNA (µg) Ratio
Diluent for DNA and Transfection Reagent
Incubation Time to Form GenJet™/DNA
Complex
Incubation time of shaved cells with transfection complex
Maximal Efficiency
Transfection
Results:
Reporter Gene
Plasmid
Efficiency (GFP %) |
DMEM with 4.5 g/L glucose, 10% FBS
95~100%
1.3x10^5 cells per square centimeter
4:1
Serum-free DMEM with 4.5 g/L glucose
12 minutes at RT
20 minutes at 37 degree
48 hours
EGFP
pEGFP-N3 (CMV promoter)
85% |
Storage Condition
Store at 4 °C. If stored properly, the product is stable for 12 months or longer
A Picture Showing Transfection Efficiency of GenJet™ Reagent on MEFs. MEFs
were grown per ATCC recommended culture medium and transfected with pEGFP-N3 by
GenJet™. The efficiency was checked 24 hours post transfection
Data Sheet
