Sigma-Aldrich
Polyethyleneimine
408727-250ML
My research involves making lentiviral vectors pseudotyped with vesicular stomatitis virus envelop glycoprotein (VSV-G). In this protocol, I transfect HEK-293T cells with 3 plasmids; one for lentiviral structural proteins, one for the genome and one for VSV-G. I have tried many different reagents to transfect cells with these and none of them are as cost-effective as PEI.
General transfection / Making lentiviral vectors
HEK-293T cells, PEI, Phosphate buffered saline, lentiviral plasmids
To transfect the cells, plate them overnight at 5x10^6 cells per 10-cm plate. At the day of transfection, mix the lentiviral plasmids in 1 ml of PBS. Add 3 ul of PEI (1mg/ml stock in water) per ug of DNA and mix by vortexing briefly. Keep at room temperature for 5-15 min. Drip gently on cells.Change medium 24 hour later and then harvest the virus 72-96 hours later.
PEI is stable (1mg/ml) at 4 degrees celcius for about a week. So make it fresh if possible
I have tried many different reagents for this protocol (Lipofectamine, Calcium phosphate, TransIT, BioT). However, PEI is hands down the cheapest of them all. It costs ~$120 for 250 ml. For each 10 cm plate, I use ~0.2 mg of PEI. This is enough to last for a lifetime of lentivirus production. On the other hand, commercial reagents cost a fortune. I have hundreds of dollars per month by using PEI to make lentiviruses.The drawback is that the titers are slightly lower, ~2-3 fold lower than BioT. But for the price, I will take it.Also, you can easily scale this up (just increase everything proportionally)
This procedure can be used for routine transfections as well.
Very very cheap; very simple protocol
Titers are a little lower than commercial reagents
Use if you want to make a lot of lentiviral vectors on a tight budget.
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