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ShRNA transfection


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#1 chlerella

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Posted 27 April 2011 - 08:32 AM

I have knocked down TFF1 gene in MCF7 breast cancer cells using DNA plasmid containing ShTFF1.
i performed western blot and there was no protein detected. after 3 more passages i performed western blot again and found a faint protein band.
its suppose to be a stable knock down but i m worried the protein may reappear completely before i finish my experiments.does this normally happen?
someone please share what you know about similar experience?

Thank u
:huh: chlerella

#2 pcrman

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Posted 01 May 2011 - 08:46 PM

Hi chlerella,

You have to bear in mind two facts:

First, RNAi knockdown even by vector is not 100% efficient; 2nd, your transfection of shRNA vector does not have 100% efficiency, which means after several passages, untranfected cell may become dominant, or transfected cell may gradually lose vector expression.




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