Expression comparision using Cp values - (Mar/07/2009 )

Hi all,

I am encountered a equation that I see recently.

After running a number of qPCR reaction, and Cp values are obtained from each reaction curve.

I recently see people using the equation of 2^(40-Cp) to calculate a "Value" which will be plotted on y-axis, and x-axis is just sample name. They say it is just a very early preliminary estimation of gene expression level, and with the assumption of putting same amount of cDNA in each well at the begining; and the 40 on the equation denotes the number of cycle that something happened to the reaction (I forgot, something related to if the reaction run to 40th cycle, then...........)

Could someone help me? Please, Thx in advance.

Microchip on Mar 7 2009, 05:39 PM said:

I think you mean the "delta delta Ct" formula, Cp is crossing point, Ct is cycle threshold. It is the same thing. Your delta delta Ct formula above assumes that the efficiency of the gene of interest is perfect 2. Then the (40-Cp) is supposedly the Ct difference between a treated and control sample (hence delta Ct). But in this case they probably only took the gene of interest, and did not compare relatively to a housekeeping gene. Why they set one of the Ct at 40 I don;t know, maybe their control is always Ct40, only their treated sample's Ct changes.

This is a very preliminary result, probably just screening the genes to see which one is really significantly expressed.

Hope this helps