Hi,

I was hoping that someone might be able to suggest a way forward with some siRNA array data I have been asked to analyse. There are 3 cell lines, each hybridised using triplicate samples onto Agilent two colour arrays:

CL, CL_knockdown and CL_scrambled.

Only one siRNA that has been used against the target.
I have analysed the data with Limma, using CL_scrambled as the ref in the design matrix. Unfortunately there are a substantial number of differentially expressed genes between the CL and CL_scrambled which will potentially confound the experiment.
Is there a recommended way to minimise the effect of potential off target effects, is this a possibility?

Possibly contrasting (CL_knockdown) - ( CL) ?
Remove the intersection of differentially expressed genes between CLvs CL_scrambled  and CL-knockdown vs CL_scrambled ?


I'd appreciate any advice that you can give.

Bryony



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