Limma - batch issues
0
0
Entering edit mode
@segal-corrinne-4150
Last seen 9.6 years ago
Hi, I am analysing single-channel miRNA microarray data: I have 4 treatments at 4 time points, mostly in triplicate, spread approximately evenly across 2 batches. Previously I have used ComBat to reduce batch effects and then used Limma and AgiMicroRna to get DE miRNAs. However, I read on the list somewhere that it is better to add Batch as a factor to the model in Limma rather than try and remove it. ### So this time I added Batch to my targets file...### FileName Treatment GErep Subject Time Compound Batch A1.txt A 1 1 hr4 v 1 A2.txt A 1 2 hr4 v 2 A3.txt A 1 3 hr4 v 2 B1.txt B 2 1 hr4 m 2 B2.txt B 2 2 hr4 m 1 B3.txt B 2 3 hr4 m 1 ... (I also have hr8, hr24, hr120 and Compound b and d) ###And did the following...### TS<-paste(phenoData$Time,phenoData$Compound,phenoData$Batch,sep=".") TS<-factor(TS,levels=c("hr4.v.1","hr4.m.1","hr4.b.1","hr4.d.1", hr4.v.2","hr4.m.2","hr4.b.2","hr4.d.2", "hr8.v.1","hr8.m.1","hr8.b.1","hr8.d.1", "hr8.v.2","hr8.m.2","hr8.b.2","hr8.d.2", "hr24.v.1","hr24.m.1","hr24.b.1","hr24.d.1", "hr24.v.2","hr24.m.2","hr24.b.2","hr24.d.2", "hr120.v.1","hr120.m.1","hr120.b.1","hr120.d.1", "hr120.v.2","hr120.m.2","hr120.b.2","hr120.d.2")) design<-model.matrix(~0+TS) fit<-lmFit(minimalSet,design) ### I now get an error message saying that coefficients are not estimable (partial NA coefficients for 260 probes). Last year I ran the same process but without batch and it worked fine. I would like to find miRNAs that have changed compared to the control at each time point, having taken into account the effect of batch. Any suggestions as to how I can get it to work? Thanks very much Corrinne (Additionally, I am doing the analysis in R development version as I would like to be able to get CI for log fc) [[alternative HTML version deleted]]
miRNA Microarray limma PROcess AgiMicroRna miRNA Microarray limma PROcess AgiMicroRna • 962 views
ADD COMMENT

Login before adding your answer.

Traffic: 866 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6