[BioC] HTqPCR problems

[Diane Guevremont]

Hi Heidi,
First thank you so much for your library HTqPCR, we have been using it a lot  successfully and it has made our data analysis so much more efficient.

However I have a problem that I can't resolve. I have tried many different approach without success and I was wondering if you could help us with any suggestions. I will try to explain what we have done to help so you can understand our problem.

Last year we did a pilot study with 21 samples and we ran TLDA with both card A and Card B,  we used your library to analyze our data  and we have identified 384 miRNAs from these 2 cards that were interesting to us and pooled them in  a customized card ("card C"; so we have 384miRNAs in common from both card A and card B) and ran another 70 samples. I have now analyze the new 70 samples with HTqPCR.
What I want to do next is where I have a problem.

I want to add the data of the first 21 samples(pilot study) to the new 70 samples (same treatment so just different biological replicates). I have successfully rbind card A and card B because they are the same samples (the 21 samples). 
I have then filtered out all the miRNAs that are not in "card C" so that I have identical list of miRNAs for both set of samples and I thought I could then cbind them so I could analyze for the same 384 miRNAs all 91 samples but I can't because it is telling me that they have different positions on the arrays.

> together<-cbind(qFilteredphase1,qFilteredphase2)
Warning message:
In cbind(deparse.level, ...) :
  Feature positions in qFilteredphase2 are not identical to qFilteredphase1


I thought I could sort what I called qFilteredphase2 and qFilteredphase1 to remove positions problem but that didn't work. I also try to not use in the command 

raw<-readCtData(files=files$File, path=path, column.info=list(feature=5,Ct=12)) 
 so remove the position in the list but it still gives me the warning message.

I was wondering if you could us any suggestion
thank you very much for your time and help
cheers
Diane


Diane Guévremont
Scientific Officer
Dept. Anatomy 
Scott 110
Office tel: 3073
Lab tel: 5792

diane.guevremont at anatomy.otago.ac.nz