transfection in NIH3T3 cells - (Feb/01/2007 )
I want to transfect my protein of interest in 3T3 cells.
Does anyone has any experience in transfecting this cell line.
Is it a easy cell line to transfect.
Which method should i use, calcium phosphate or fugene.
I have no experience with this cell line.
If i were to try, i would start with calcium phosphate (much cheaper) and if this doesnot work, I would try fugene or lipofectamine.
I just saw the topic, so sorry for the late answer 
My colleague has a lot of experience transfecting NIH3T3 cells. He uses Fugene at a ratio of 3:1 (Fugen:DNA). It seems to be a pretty easy cell line to transfect!
I transfect 3T3 L1 cells using Transfast (Promega), efficiency is not wonderful but more than enought for luciferase, I don't know about other uses. IIRC I got a similar efficiency using Superfect (Qiagen).
I've been working with 3T3-L1 cells for a while, and I have never had any good experiences with transfection. So I use retroviruses or lentiviruses, and I have an excellent efficiency (over 90%).
Hi,
I used retroviral vector to transfect NIH/3T3 cells. It is very easy to transfect these cells using retrovirus.
Sometimes, I even think the expression level of protien of interest is too high. Maybe you can choose inducible retroviral vector. Good lucks!
I used retroviral vector to transfect NIH/3T3 cells. It is very easy to transfect these cells using retrovirus.
Sometimes, I even think the expression level of protien of interest is too high. Maybe you can choose inducible retroviral vector. Good lucks!
Do u think electroporation'll work for 3T3-L1 cells?
I've no experience with retro & lenti viruses.
Can u please give me an idea that how it works?
Retrovirus are pretty simple. You need to clone you gene into a retroviral vector, and co-transfect the obtained vector with a envelope coding plasmid into a packaging cell line (i.e. 293T). The viruses will be released in the supernatant, and is ready to use.
Now, I don't use retrovirus for surexpression, so I don't know about plasmids that can do surexpression. cvdrchan surely does know.
You can check on the clontech website. They have a pretty complete pdf document about retroviral gene transfert.
Hope this helps!
I've tried calcium phospahte and electroporation but no luck.Few cells were transfected by Lipofectamine plus but it was not too good.
Does someone has any experience in transfecting these cells.
Plz help.What should i do?
Can someone give me any protocol for transfecting this cell line.