Protocol needed stain Cos cells for DNA,RNA

m-brazil at nimr.mrc.ac.uk m-brazil at nimr.mrc.ac.uk
Tue Jul 11 07:16:47 EST 1995


> 
> We have transfected by electroporation, Cos-1 cells with an expression 
> vector containing a retroviral gene.  Is there a way to probe cells on a 
> slide for DNA and RNA.  We want to determine if the cells have been 
> transfected and if there is RNA being made from the construct.  I know we 
> could isolate RNA and DNA and do Southerns and NOrtherns but, we are 
> looking for something quicker and in vitro.  Thanks for any help.  
> Regards, Mary
> 
> 
> 
Dear Mary,
I use PCR to check for positive transfectants. Lyse 1x105 cells from a particular clone and just add a small amount to your standard PCR reaction.

You obviously need primers specific for your transfected DNA. A positive PCR however doesn't mean that you will have expression, as integration may have occured somewhere else in your DNA that isn't amplified by the primers.

If you want more details let me know.

Good luck
Melanie
m-brazil at nimr.mrc.ac.uk
> 




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