Description
GenJet™ DNA In Vitro Tranfection Reagent for HuH-7 Cells is pre-optimized for human HuH-7 cell transfection. HuH-7, derived from human cancer liver, is very hard to transfect. With our pripariotary peptide conjugation technology, this reagent was formulated to transfect HuH-7 cells with up to 70% efficiency (GFP positive cells).
Refer to the following optimal transfection conditions for maximal transfection efficiency on HuH-7 cells. GenJet™ reagent, 1.0 ml, is sufficient for 300 to 600 transfections in 24 well plates or 150 to 300 transfections in 6 well plates.
Summary of Optimal Transfection Conditions:
Confluence on the day of transfection
|
90% |
| Cell culture conditions |
DMEM with 4.5 g/L glucose, 10% FBS
|
| GenJet™ (µl) : DNA (µg) Ratio |
3:1 |
Diluent for DNA and Transfection Reagent
|
Serum-free DMEM with 4.5 g/L glucose
|
| Incubation Time to Form GenJet™/DNA Complex |
15 minutes at RT |
| Presence of Serum/Antibiotics during Transfection |
Yes |
| Change Medium After Transfection |
Yes, ~5 Hours After Transfection |
| Maximal Efficiency |
48 hours |
Transfection Results:
Reporter Gene
|
EGFP |
Plasmid
|
pEGFP-N3 (CMV promoter)
|
| Efficiency (GFP %) |
80% |
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 HuH-7 Cells
GenJet™ reagent is optimized for
HuH-7 cells with
exceptional efficiency in comparison of Lipofectamine 2000 (L2K) and Fugene HD.
HuH-7 cells
were grown per ATCC recommended culture medium and transfected with pEGFP-N3 by
GenJet™. The efficiency was checked 48 hours post transfection
Right Panel: Comparison of transfection efficiency of GenJet with Lipofecatmine 2000 (L2K), and Fugene
HD on HuH-7 cells. GFP DNA (pEGFP-N3) was introduced to HuH-7 cells with different transfection reagents per manufacturer’s protocols. GFP positive cell (%)
and fluorescence intensity were detected by passing through FACS 48 hours post transfection
Left Panel: presence of serum and antibiotics enhances GenJet
efficiency on HuH-7 cells. HuH-7 cell (grown on collagen treated dishes)
was transfected with three different conditions——-serum and antibiotics
free, presence of 10% serum and antibiotics followed by removal 5 hours post transfection and presence of 10% serum and antibiotics without removal 5 hours
post transfection.
GenJet™ reagent is optimized for
HuH-7 cells.
HuH-7 cells
were grown per ATCC recommended culture medium on a collagen type I treated
culture dish and transfected with GFP cDNA (pEGFP-N3) by GenJet™. The efficiency was checked
24 hours post transfection
by Nikon Elipse 2000 confocal microscopy
Data Sheet
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