Hi,

The common way is to use the Ct values to 'relative gene expression' compared to some reference genes. The other way is to do 'absolute quantification'. Both the approaches are enabled in the software application of most instruments. See what is appropriate for your analysis

Hi,

Plotting directly ct value has become an old and outdated way. You need to follow livak formula to assess the obtained ct value. Then either ddct (relative expression) or 2ddct/2^ddct (fold change) calculated value which will come that should be plotted for the representation.

Regards

Saurabh

@ Lakshani Perera

I would like to add some more information to what suggested by Saurabh Mandal

The 2-ΔΔCq (Livak) Method which assumes that both target and reference genes are amplified with efficiencies near 100% and within 5% of each other. Before using the Livak method, it is essential to verify these assumptions by determining the amplification efficiencies of the target and the reference genes.

The ΔCq Method Using a Reference Gene is a variation of the Livak method that is simpler to perform and gives essentially the same results. In contrast to the ΔCq values obtained with relative quantification normalized against a unit mass, this method uses the difference between reference and target Cq values for each sample.

The Pfaffl Method

The Livak method for calculating relative gene expression is valid only when the amplification efficiencies of the target and reference genes are similar. If the amplification efficiencies of the two PCR products are not the same, an alternative formula must be used to determine the relative expression of the target gene in different samples.

It is the experimental setup, which decides the needed representation and formula to be used.

For more details visit

https://www.bio-rad.com/en-id/applications-technologies/real-time-pcr-experimental-design?ID=LUSNJVIVK