Hello,
Some references have mentioned the following equations for T/S ratio?
The Relative telomere length from each sample, based on the T/S ratio, was calculated as the ∆Ct test (Ct(telomere)−Ct(beta-globin)) normalized to the average T/S ratio of the three standard DNA samples on the same plate ∆Ct ref (Ct(telomere)−Ct(beta-globin)). Hence, the final equation was 2− (ΔCt test −∆Ct ref)=2−∆∆Ct. (doi:10.1136/bmjopen-2017-020263 )
or
Telomere/single-copy gene (T/S) values were calculated by the formula 2-ΔCt where ΔCt =Ct telomere – Ctβ2globin. while Relative T/S values were determined by dividing sample T/S values with the T/S value of reference DNA .(www.pnas.orgcgidoi10.1073pnas.0501724102)
or
The relative TL was the quotient of the average quantity of TL and HBB.
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Consider the following case-
If the standards for T and S amplicon range from 8ng/ul to 0.5 ng/ul and used only for checking PCR efficiency. While the sample under analysis if we used 10 ng DNA (2.5ng/ul) then
Can we calculate the T/S ratio as follows?
Mean of 3 Ct values of T amplicon divided by Mean of 3 Ct values of S amplicon
( Mean of Ct for T/ Mean of Ct for S)
Please suggest.
Hi colleague
Hope the following paper may help you:
http://depts.washington.edu/risques/wordpress/wp-content/uploads/2015/09/Cawthon-2002.pdf
Regards....
Thanks for sharing your opinion and references. But actually my concern is about the method used for getting T/S ratio and Relative T/S ratio
There are different ways quoted in references for T/S and Relative T/S ratio calculation like as I mentioned in my first question.
1) Some are using subtraction and division combinations
2) Some are using the only subtraction for T/S and Relative T/S
3) Some are using division and then subtraction combination
4) Some are using division only for T/S and relative T/S
So which is the correct way?
Can we calculate the T/S ratio and Relative T/S ratio as follows?
First, calculate the T/S ratio by
Mean of 3 Ct values of T amplicon divided by Mean of 3 Ct values of S amplicon
( Mean of Ct for T/ Mean of Ct for S)
Then calculate the relative T/S ratio by
Relative T/S ratio= T/S ratio of the test sample divided by the T/S ratio of the reference sample
Please suggest.
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