SiRNA transfection in HEK293T - (May/03/2005 )

Hi everyone,

I'm a new user of this forum...and in this field of research as well !
I have to set a gene silencing experiment with SiRNA in HEK293T cells.
I want to perform it in 24-wells plate and my question is : is there anyone who use these cells and what density was the cells plated at ? and for how long ?
Thank you for your answers in advance

Best regards

David

Hey Bulliwyf,

We have transfected both plasmid and siRNA into HEK293 cells using the calcium phosphate method. For this we tend to plate the cells out 24 hours before transfection at a density of 1x10(5) cells/ml in a total of 5mls in 60mm dishes...I guess you would scale down for your 24 well plates. Cells are harvested and RNA and protein levels analysed 24-48 hours after transfection, I have done both time points and both have given comparable results but for consistency I tend to stick to 48hours!

Hope that is of some help

holly