5 replies to this topic

[cardosopedro]

Hello.

I'm analyzing the expression of one gene after induction of cell death with a drug. I am using a wild type strain as well as a knockout mutant strain (for a cell signaling protein, in order to check if the activation of the gene was dependent on this signaling pathway). For the analysis I use the wild type control-treated sample as a calibrator, meaning the expression is 1 in this sample. I have these levels of expression:

wild type control: 1
wild type drug: 50
mutant control: 10
mutant drug: 50

In the mutant I also have induction of the gene to the same levels as in the wild type. But if I consider that in the mutant control sample I already have 10 times more expression of the gene than in the wild type, the induction caused by the drug is only 5 (50/10 = 5) whereas in the wild type this is 50 (50/1 = 50).

Should I consider that the mutant has a decreased capacity to induce the gene or not?


Please leave your comments.
Thanks.

[ttt1]

Your mutant could be deficient in a repressor of gene expression that inhibits action of a separate activating pathway- removing something that keeps gene expression at a basal level might lead to the pattern you're observing.

Edited by ttt1, 22 April 2012 - 06:03 PM.

[cardosopedro]

Thanks for the answer. Yep, that is one possibility.

I don't know if I can conclude that the difference that I observe in the drug-induced expression in the mutant means that the induction of the gene is impaired in this strain... because although there is a significative difference between drug-induced fold change between wild type and mutant, the truth is after the drug the expression reaches the same levels as in the wild type...

[Do1ngP3e]

Are you doing absolute quantitation or relative?

[cardosopedro]

Relative.

[cardosopedro]

Any comments?