Relative quantification

Relative Quantification Data Presentation

How to present the ratio data?In gene expression analyses qPCR is most commonly used for relative quantification, meaning that we are comparing one sample to another, treatment to control, one time point to another etc. After we calculate the ratio between the selected sample(s), using any of the available methods, we have to face another challenge: how to present the ratio data?

Let’s take a look at the example of gene expression of one gene in 5 samples, compared to control. Samples A and B are up-regulated, we did not detect any change in sample C, while samples D and E were strongly down-regulated. While ratio provides an intuitive measure of expression changes, the up-regulation and down-regulation are not presented symmetrically. We can clearly see the difference between up-regulated samples A and B, but we cannot distinguish that the down-regulated samples D and E are actually 20-fold different. Asymmetry of those values will also affect statistical testing (See Picture 1).

Sample-Ratio-1

The most widely used alternative transformation of the ratio is the logarithm base 2, which has the advantage of producing symmetrical values and treating up- and down-regulated genes in a similar way . If we look at log2 values of relative expression, ratio of 1 (sample C) becomes 0, which conveniently means there is no expression change and values of down-regulated samples (C and D) become negative. From this kind of presentation it is easier to see the differences between samples C and D (See Picture 2).

Sample - Ratio-2

 

By Spela Baebler, PhD, Research Associate, Department of Biotechnology and Systems Biology, National Institute of Biology, SI





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