variation in transfection efficiency

guo at mcclb0.med.nyu.edu guo at mcclb0.med.nyu.edu
Mon Apr 19 00:50:44 EST 1993


In article <1993Apr16.155627.11171 at gserv1.dl.ac.uk>, ff at rri.sari.ac.uk (Fiona Fairhurst) writes:
> Can anyone explain why we get such huge variation in transfection
> efficiency efficiency when using the calcium phosphate method to
> transiently transfect C2C12 myoblasts with a CAT plasmid?  We have tried
> numerous theories ranging from pH to amount of DNA to length of
> precipitate formation but still cannot explain why on one day
> transfection is high and on another low.  Has anyone else experienced
> this? or does anyone have suggestions of how to improve consistancy?

Fiona: Hi.  In my experience with other cell lines, the problem with
inconsistant transfection efficiency are mainly due to the various time I let 
my DNA-CaPO4 ppt sit before adding to the cells. If I add DNA-CaPO4 ppt to
cells immediately (rather than sitting for 20 min,as suggested in the "Bible")
I get consistant, and high efficiency of transfection.
   I hope this helps,
   
Sheng Guo
NYU Medical center



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