Dear all,
I am wondering what is the best way to compare gene expressions in Taqman qPCR. Is it the 2^ddCT values or the 2^dCT?
I want to compare the relative expression of different assays in 2 populations: wild type versus knockout mice.
Thanks a lot in advance!
Laurence Stuart Dawkins-Hall
Popular answer
Dear Alaa
- I am the above person referred to in the previous answerr (so thank you Saurabh !!)
- You should absolutely use 2^ ddCt not 2^ct in which I assume you would be just be comparing the difference in Ct value between your WT and mutant mouse; This would not normalise against a housekeeper which you absolutely need to do when comparing 2 different cDNA sources (biological replicates)
- Even so, the 2^DDCt method is really for comparing the same model treated and intreated not strictly speaking 2 different systems (mice)
- You woiuld be better determining absolute expression by standard curve in the 2 different systems and then computing the quotient to obtain expression difference
- Take a look at the guide I have attached for absolute expression (using standard curves)
3 votes
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Saurabh Mandal
You can go through the answer of two people, they explain nicely in context of qPCR data analysis.
https://www.researchgate.net/profile/Laurence_Dawkins-Hall
Jochen Wilhelm
1 votes
1 thanks
Laurence Stuart Dawkins-Hall
Dear Alaa
- I am the above person referred to in the previous answerr (so thank you Saurabh !!)
- You should absolutely use 2^ ddCt not 2^ct in which I assume you would be just be comparing the difference in Ct value between your WT and mutant mouse; This would not normalise against a housekeeper which you absolutely need to do when comparing 2 different cDNA sources (biological replicates)
- Even so, the 2^DDCt method is really for comparing the same model treated and intreated not strictly speaking 2 different systems (mice)
- You woiuld be better determining absolute expression by standard curve in the 2 different systems and then computing the quotient to obtain expression difference
- Take a look at the guide I have attached for absolute expression (using standard curves)
3 votes
3 thanks
Alaa Ibrahim
Dear Laurence!
Thanks a lot for your very very kind reply!! I appreciate it very much! I took a look at the PDF and am following it!
All the best to you! Thanks again
Alaa
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