Transfection

brett brett at BORCIM.WUSTL.EDU
Thu Feb 13 23:51:29 EST 1997


>Fellow netters,
>
>I'm want to find which method of transfection results in the highest
>percentage of cells taking up DNA. I want to do a transient transfection,
>followed by a western. At present my western signal is very weak if present
>at all. I think this is due to only a small number of cells being
>transfected. I am presently using chicken embryo fibroblasts.
>
>Your thoughts on this would be helpful.
>
>Steve 

We've had good luck electroporating primary CEF's with RNA. I imagine you could
adapt these conditions for DNA:
0.2mm gap cuvette
0.5ml cells in PBS (we use RNase-free), 2x10e7/ml
BioRad GenePulser settings:
        25uF
        infinite resistance
        1.5kV



Brett Lindenbach
    
Program in Immunology                              
Washington University - St Louis                  
brett at borcim.wustl.edu                             




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