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 [[alternative HTML version deleted]]

Bryony from what I understand from your mail, your question has nothing to do with off-target effects. And the first thing you should do at this point is talk to the person who designed and performed the experiment. Best wishes Wolfgang On 28/09/10 23:52, Lloyd, Bryony wrote: > 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 > > > > [[alternative HTML version deleted]] > > _______________________________________________ > Bioconductor mailing list > Bioconductor at stat.math.ethz.ch > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor -- Wolfgang Huber EMBL http://www.embl.de/research/units/genome_biology/huber