Hello,
I need to induce EMT by TGFb recombinant Protein, and also I want to transfect the X cell line with my miRNA candidate and inhibitor.
I need an efficient protocol please!
First inducing with TGFb ( how long)
Then transfection with miR - mimics!
any Recommendation would highly be appreciated
Thanks in Advance
Dharmacon mimics and inhibitors work well although Exiqon inhibitors work well also.
Mimic final concetrations around 30 nM should yield a few hundred-fold higher levels of your miRNA of choice. Inhibitors are trickier and typically require a dose response experiment. Typically I have used 30-90 nM of various miRNA inhibitors to get a decent functional effect.
Not sure which transfection reagent works best with your cell line but they all work fairly well. If transfection efficiency is an issue, try transfecting in suspension immediately before plating your cells.
Thanks Rozan,
I already use it for mimics transfection but I need an efficient protocol for inducing EMT by TGFb recombinant protein.
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