# DATA ANALYSIS for qPCR - Calculating 2^(-Del Del Ct). (Jan/19/2010 )

Hello,

I am trying to analyze data using the 2^(-DDCt) method and I had a question. I run my samples in triplicate. so while doing the calculations, I calculate the Delta Ct (target-HKG) and then Del Del Ct value. Then I calculated the 2^(-Del Del Ct) values for each sample and then averaged the values. I also calculated the 2^(-Del Del Ct) value by first averaging the Del Del Ct value (alternate way).

When I do this I get different values for my 2^(-Del Del Ct) (averaged) and hence different results. Which is the correct way to calculate 2^(-del del Ct) and analyze the results.

Thanks

Pmaj

-pmaj-

pmaj on Jan 20 2010, 05:12 AM said:

Hello,

I am trying to analyze data using the 2^(-DDCt) method and I had a question. I run my samples in triplicate. so while doing the calculations, I calculate the Delta Ct (target-HKG) and then Del Del Ct value. Then I calculated the 2^(-Del Del Ct) values for each sample and then averaged the values. I also calculated the 2^(-Del Del Ct) value by first averaging the Del Del Ct value (alternate way).

When I do this I get different values for my 2^(-Del Del Ct) (averaged) and hence different results. Which is the correct way to calculate 2^(-del del Ct) and analyze the results.

Thanks

Pmaj

i average the Ct values of my triplicates, then determine the delCt values (ie gene of interest - constitutive gene) of sample and control separately. Then find out the deldel Ct (mutant delCt - control delCt) .....use this value to calculate the fold change, i.e, 2^(-deldel Ct).....hope this helps....

-DRN-

DRN on Jan 20 2010, 03:42 AM said:

pmaj on Jan 20 2010, 05:12 AM said:

Hello,

I am trying to analyze data using the 2^(-DDCt) method and I had a question. I run my samples in triplicate. so while doing the calculations, I calculate the Delta Ct (target-HKG) and then Del Del Ct value. Then I calculated the 2^(-Del Del Ct) values for each sample and then averaged the values. I also calculated the 2^(-Del Del Ct) value by first averaging the Del Del Ct value (alternate way).

When I do this I get different values for my 2^(-Del Del Ct) (averaged) and hence different results. Which is the correct way to calculate 2^(-del del Ct) and analyze the results.

Thanks

Pmaj

i average the Ct values of my triplicates, then determine the delCt values (ie gene of interest - constitutive gene) of sample and control separately. Then find out the deldel Ct (mutant delCt - control delCt) .....use this value to calculate the fold change, i.e, 2^(-deldel Ct).....hope this helps....

Thanks,

I am going to try that and see how that works.

Bests,

Pmaj

-pmaj-