[BioC] contrast.fit and different p-values for same comparison

[Naomi Altman]

The point of eBayes is to use all of the available data.  If you use 
only 18 arrays, the available data are only these arrays, so your 
results differ.  Even if you do not use eBayes, the MSE used for the 
statistical tests will depend on the arrays included in the analysis.

Regards,
Naomi

At 02:17 PM 2/12/2010, Lakshmanan Iyer wrote:
>Hi
>I used limma... topTable on the "same contrast" for a complete dataset
>(36 arrays, 12 conditions, 3 replicates per condition) and a subset of
>the same (18 arrays).
>I used lmFit, followed by contrasts.fit and eBayes in both cases.
>
>However, I get different Ave Expression and P-values in the two cases.
>
>Why is this happening? I cannot readily explain this.
>
>Thanks for any help/clues.
>
>The first row is full dataset
>second row is the subset
>
> > rbind (x[x[,1]=="ILMN_2772632",],x1[x1[,1]=="ILMN_2772632",])
>                  ID    logFC  AveExpr        t      P.Value    adj.P.Val
>37561  ILMN_2772632 4.198562 10.40451 22.86259 2.007820e-21 9.091611e-17
>375611 ILMN_2772632 4.198562  9.85615 24.89322 1.224698e-17 1.631045e-14
>               B
>37561  37.44148
>375611 29.95781
>----------------------------------------
>-best
>-Lax
>Here is the snippet of code with some output.
>
>library (limma);
>eset <- read.delim("results/expressions.tsv",sep="\t",stringsAsFactors=F)
>rownames(eset) <- eset[,1]
>eset <- eset[,-1]
>names(eset) <- gsub("^X","",names(eset))
>samples <- read.delim("mySamples.tsv",sep="\t",stringsAsFactors=F)
>TS <- factor(samples$Condition, levels=unique(samples$Condition))
>design <- model.matrix(~0 + TS)
>colnames(design) <- levels(TS)
>cont.matrix <- makeContrasts(Day1AgedPnxVsCtrl = Day1AgedPnx -
>Day1AgedControl, Day3AgedPnxVsCtrl = Day3AgedPnx - Day3AgedControl,
>Day7AgedVsCtrl = Day7AgedPnx - Day7AgedControl, Day1YoungPnxVsCtrl =
>Day1YoungPnx - Day1YoungControl, Day3YoungPnxVsCtrl = Day3YoungPnx -
>Day3YoungControl, Day7YoungPnxVsCtrl =Day7YoungPnx - Day7YoungControl,
>levels=design)
>
>fit <- lmFit(eset, design)
>fit2 <- contrasts.fit(fit,cont.matrix)
>fit2 <- eBayes(fit2)
>x <- topTable(fit2, coef="Day1AgedPnxVsCtrl",num=dim(eset)[1])
>#
>TS1 <- factor(samples$Condition[1:18], levels=unique(samples$Condition[1:18]))
>design1 <- model.matrix(~0 + TS1)
>colnames(design1) <- levels(TS1)
>cont.matrix1 <- makeContrasts(Day1AgedPnxVsCtrl = Day1AgedPnx -
>Day1AgedControl, Day1YoungPnxVsCtrl = Day1YoungPnx - Day1YoungControl,
>levels=design1)
>fit1 <- lmFit(eset[,1:18], design1)
>fit21 <- contrasts.fit(fit1,cont.matrix1)
>fit21 <- eBayes(fit21)
>topTable(fit21, coef="Day1AgedPnxVsCtrl")
>x1 <- topTable(fit21, coef="Day1AgedPnxVsCtrl",num=dim(eset)[1])
>
> > dim (eset)
>[1] 45281    36
> > dim (design)
>[1] 36 12
> > dim (cont.matrix)
>[1] 12  6
> > dim (design1)
>[1] 18  6
> > dim (cont.matrix1)
>[1] 6 2
> > rbind (x[x[,1]=="ILMN_2772632",],x1[x1[,1]=="ILMN_2772632",])
>                  ID    logFC  AveExpr        t      P.Value    adj.P.Val
>37561  ILMN_2772632 4.198562 10.40451 22.86259 2.007820e-21 9.091611e-17
>375611 ILMN_2772632 4.198562  9.85615 24.89322 1.224698e-17 1.631045e-14
>               B
>37561  37.44148
>375611 29.95781
>
>
>-------------
> > sessionInfo()
>R version 2.10.1 (2009-12-14)
>i486-pc-linux-gnu
>
>locale:
>  [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C
>  [3] LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8
>  [5] LC_MONETARY=C              LC_MESSAGES=en_US.UTF-8
>  [7] LC_PAPER=en_US.UTF-8       LC_NAME=C
>  [9] LC_ADDRESS=C               LC_TELEPHONE=C
>[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
>
>attached base packages:
>[1] stats     graphics  grDevices utils     datasets  methods   base
>
>other attached packages:
>[1] limma_3.2.1
>
>loaded via a namespace (and not attached):
>[1] tools_2.10.1
> >
>
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Naomi S. Altman                                814-865-3791 (voice)
Associate Professor
Dept. of Statistics                              814-863-7114 (fax)
Penn State University                         814-865-1348 (Statistics)
University Park, PA 16802-2111