Antibiotic after transfection or not? - (Aug/16/2006 )
I think this is a very stupid question, but I feel I'm missing something here.
I've only worked with bacteria, fungi and yeast and now I have to transfect cells.
I'm working with the pCDNA3.1 vector and I will probably use the FuGENE6 for transfection.
But according to the protocols, you add the transfection mix and that's it. You don't add any antibiotic. They only talk about about antibiotica when you want to make a stable cell-line. Is this true?
I don't want to make a stable cell-line, I just want to transient overexpress a couple of genes and see the effect. But isn't it necessary to select transformants with antibiotics just like in coli's?
I feel so stupid at this moment!
I am also using FuGene6 for transfecting and we don't use an extra antibiotic. You just tranfect the cells and do a parallel transfection with GFP to see your transfection efficiency.
Your overexpressed genes will make an effect, even if you don't have a 100% transfection rate.
I have a very poorly transfactable cell line, which only shows around 30% transfection efficiency with GFP and I see a difference for my transfected genes. The amount of expressed protein is sufficiently high to change signal transduction, which I am looking for. We only use G418 for the stable transfection, as you mentioned.
if you want to select transformants , you use antibiotics to kill non transfected cells, and then you are on the road of the stable cell line.
In the transient transfection, you transfect, you pray any god that you hev enough transfected cells and you do your assay in the next 24 hours. 24 hours is too short to do any selection. you need several days, or several weeks in any way.