ShRNA transfection - (Apr/27/2011 )
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?
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.