RT-PCR performed in gene silencing but mechanistically what are the benefits of this procedure?


closed as unclear what you're asking by AliceD, March Ho, The Last Word, kmm, fileunderwater Dec 7 '15 at 11:40

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    $\begingroup$ Can you please re-phrase your question? At the moment it is a bit unclear what you want to know exacty. $\endgroup$ – Chris Dec 2 '15 at 17:47

RT-qPCR amplifies mRNA molecules present in a cell, if you have the right primers for it. The whole point of RNAi (the "gene silencing" that I think you're referring to) is downregulating the expression of a specific gene, so you'd expect the mRNA transcript of said gene to be not present (rather, present at very low concentrations at any given timepoint) if indeed the interference has worked.

Therefore, after RNAi has been performed, you can check by qPCR that there are indeed very low levels of the specific mRNA present in the cell. You might want to also perform a Western Blot against control cells transfected with scramble siRNA (where nothing should be downregulated, think of it as an "empty" siRNA), to check that protein levels are indeed much lower than in "regular" cells.


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