[BioC] Interpreting topTable results for limma factorial design

[James W. MacDonald]

Hi Sally,

You aren't going to be able to get an Excel attachment through the 
listserv (it will be stripped). Your best bet is to put the data inline.

Best,

Jim



Sally wrote:
> I have a 2x2 factorial design which I ran through limma.  The factors are (1) species  [(Coho (c)) and Sockeye (s))] and (2) sample time (0, 24, 48 and 96 hours).
> 
>  
> 
> The R script used was:
> 
>  
> 
> source("http://bioconductor.org/biocLite.R")
> 
> library(limma)
> 
> library(Biobase)
> 
> exprdata<-read.table("exprsData.txt", header=TRUE,sep="\t",row.names=1,as.is=TRUE,fill=TRUE,)
> 
> phenotypicdata<-read.table("phenotypicdata.txt",row.names=1,header=TRUE,sep="\t")
> 
> myexprdata<-as.matrix(exprdata)
> 
> myphenotypicdata<-as.data.frame(phenotypicdata)
> 
> adf<-new("AnnotatedDataFrame",data=phenotypicdata)
> 
> eset<-new("ExpressionSet",exprs=myexprdata,phenoData=adf)
> 
> targets <- readTargets("targets.txt")
> 
> TS <- paste(targets$Species, targets$Time, sep=".")
> 
> TS <- factor(TS)
> 
> design <- model.matrix(~0+TS)
> 
> colnames(design) <- levels(TS)
> 
> fit <- lmFit(eset, design)
> 
> cont.matrix<-makeContrasts(s0vss24=s.0-s.24, s24vss48=s.24-s.48, s48vss96=s.48-s.96, c0vsc24=c.0-c.24, c24vsc48=c.24-c.48, c48vsc96=c.48-c.96, s0vsc0=s.0-c.0, s24vsc24=s.24-c.24, s48vsc48=s.48-c.48, s96vsc96=s.96-c.96, levels=design)
> 
> fit2 <- contrasts.fit(fit, cont.matrix)
> 
> fit2 <- eBayes(fit2)
> 
> c48vsc96<-topTable(fit2,coef="c48vsc96",number=400,adjust.method="BH",p.value=1)
> 
>  
> 
> I have appended an Excel file which includes both the topTable results and the average M values for both coho 48 hours and coho 96 hours for those genes with a p value less than 0.05.
> 
>  
> 
> My questions are:
> 
>  
> 
> For gene #1, what does the logFC mean?  
> 
> At which sample time (48 or 96 hours) is this gene down-regulated -2.67 (fold change)?
> 
> The logFC does not appear to correlate with either of the two M values (48hrs = +0.63, 96 hrs = +2.42)?
> 
> Something seems wrong to me.
> 
>  
> 
> Thanks 
> 
>  
> 
> Sally Goldes
> 
> 
> 
> 
> 
> 
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-- 
James W. MacDonald, M.S.
Biostatistician
Affymetrix and cDNA Microarray Core
University of Michigan Cancer Center
1500 E. Medical Center Drive
7410 CCGC
Ann Arbor MI 48109
734-647-5623