I am co-transforming two plasmids (both 8kb) into HEK cells using Lipofectamine 2000, followed by infection, but each time the efficiency of transfection is very low (5-10%) and the more cell death . what should I do? please suggest. is it might be caused by the large size of the plasmids?
If 8kb is the full size of the plasmid, it is not too big. What kind of HEK cells are you using? HEK293T would have better transfection efficiency than HEK293. For co-transfection, I recommend using reverse transfection. I typically get >90% transfection efficiency and >95% co-transfection efficiency.
thank you for your feed back, I am using HEK293 Tcells, both plasimds are 8 KB (total 16KB). I will try to apply reverse transfection. Hope it goes well.
Hi Muhammad,
Maybe your protein is just toxic so your efficiency is low because most cells die?
-ED
I transfect HEK293T cells all the time with lentiviral vectors (>8Kbp) without a problem with inefficiencies higher than 90%. Stay away from Lipo2000. Too toxic. Use JetPrime, cheaper, more gentle, and higher efficiency.
I can send you a protocol and everything if you need
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