2-ΔΔCT interpretation of gene copy number

[piano6179]

Hello,

I'm trying to see if my gene has an increased copy number in tumor A and tumor B. I used normal tissue as a calibrator which gave a ΔΔCT value of 0 and therefore the 2-ΔΔCT is 1 which is great.

2-ΔΔCT (2 to the power of minus Delta Delta CT)

However, when I calculated the 2-ΔΔCT (which indicates the copy number) for my gene, in tumor A it gave a value of 1.84 and in tumor B, the 2-ΔΔCT was 0.7.

Does anyone know how to interpret these data? Am I missing something?

Shall I be mutiplying the 2-ΔΔCT value of my normal tisse (calibrator) by 2 since it has 2 alleles? and therefore multiply the rest of the 2-ΔΔCT vlaues by 2? or does the 2-ΔΔCT take that into consideration?

I guess my question is: what does 1.84 mean? that the gene has almost 2 copies only in tumor A? Isn't it normal to have 2 copies of the gene anyway? If so then what does 0.7 mean?

Any clue? I'm totally confused :s

Thanks alot

[amicia]

Hi,
values under 1 using 2-ΔΔCT mean that the gene is underexpressed, while on contrary, values upper to 1 mean that the gene is overexpressed. The problem with the underexpressed genes is that tehy vary between o and 1, while the overexpressed between 1 to infinite. Shall I suggest You applying to the results with a base 2 log, in order to trasform the results between - infinite and + infinite (obviously 0 mean not changes)?

[piano6179]

Hi Amicia,

Thanks alot. I'll do that. It turned out that the 2-ΔΔCT value I get is for haploid cells so you have to mutiply it by 2 or use this: 2(1+E)^-ΔΔCT (where E is the PCR Efficiency close to 100% so E=1).

It makes sense now that the 2-ΔΔCT for the normal calibrator is 1 which means it has 2 copies of the gene when multiplying it by 2 or using the above equation.