normalize the data in respect to the housekeeping gene in R / how to get the Ct values from the raw data

Question

[anm17]

Hello community,

I am very new to R and gene expression Analysis, so i hope my question does not sound that silly

So I am trying to reproduce the results of a certain Series from the GEO database. According to their paper they normalized the data in expression level of a certain housekeeping gene.

I have the .cel files and created a phenodata file, with the name, filename and target information and saved the data with the function readaffy in a variable. So i am not sure how to normalize the data in respect to the housekeeping gene in R. (i know how to do it in excel, but could not find an option to open the cel files in excel).

Another question is: they used the ddCT method to calculate the fold induction value, but to recalculate it with this method I would need the Ct , am I right? Is there a way to get it through the raw dat (.cel files)?

Thank you very much!

Answer 1

[Benn]

It seems like you are confusing two things now, cel files are from affymetrix and are microarrays. Ct values come from qPCR. Both methods can be used to measure gene expression, but both require different approaches of data analysis. In other words, you don't get Ct values from microarray data.

Answer 2

[anm17]

@b.nota Thank you ! This makes sense for me now, but does it mean if they don't provide the Ct values, there is no option for me to reproduce their results?

[Benn]

Most qPCR data are not deposited in databases such as GEO. You can ask the authors for the raw qPCR data, I am not sure if they want to share it...

[anm17]

Oh i see, thank you very much!