I have a WT sample from which we have sorted various cell populations via FACS. As a control population, we usually sort a pool of mixed cells from the same sample. Using RT-qPCR, I simply want to quantify the expression of marker genes A and B with respect to a housekeeping gene - in each targeted population and the control mixed population. Can the delta Ct value, say for gene A in population X, be directly used as an expression value for gene A in population X?
Hi Annasha,
To the best of my understanding, delta Ct value (Ct gene A - Ct housekeeping gene) is not an absolute measurement for gene expression of gene A but rather a relative expression in fold change compared to your housekeeping gene, you need to use 2^(-delta delta Ct value) as a relative expression value where you normalize to your control mixed population. You will then plot the 2^(-delta delta Ct value) as the y-axis of your results. This means that compared to control samples, gene A is expressed Y fold more or less than housekeeping genes, and you can use this fold change to directly compare among multiple populations you sorted.
Best,
Zhixin
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