# Version info: R 2.14.1, Biobase 2.15.3, GEOquery 2.23.2, limma 3.10.1
# R scripts generated  Tue Dec 22 16:00:35 EST 2015
## This script is archived at http://oncinfo.org/file/history/Geo2R.REDO.R
#################################################################################
#   Differential expression analysis with limma
library(Biobase)
library(GEOquery)
library(limma)
# load series and platform data from GEO
gset <- getGEO("GSE15061", GSEMatrix =TRUE)
if (length(gset) > 1) idx <- grep("GPL570", attr(gset, "names")) else idx <- 1
gset <- gset[[idx]]
# make proper column names to match toptable 
fvarLabels(gset) <- make.names(fvarLabels(gset))
# group names for all samples
sml <- c("G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G0","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","G1","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","G0","G0","G0","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X","X");
# eliminate samples marked as "X"
sel <- which(sml != "X")
sml <- sml[sel]
gset <- gset[ ,sel]
# log2 transform
ex <- exprs(gset)
qx <- as.numeric(quantile(ex, c(0., 0.25, 0.5, 0.75, 0.99, 1.0), na.rm=T))
LogC <- (qx[5] > 100) ||
          (qx[6]-qx[1] > 50 && qx[2] > 0) ||
          (qx[2] > 0 && qx[2] < 1 && qx[4] > 1 && qx[4] < 2)
if (LogC) { ex[which(ex <= 0)] <- NaN
  exprs(gset) <- log2(ex) }
# set up the data and proceed with analysis
fl <- as.factor(sml)
gset$description <- fl
design <- model.matrix(~ description + 0, gset)
colnames(design) <- levels(fl)
fit <- lmFit(gset, design)
cont.matrix <- makeContrasts(G1-G0, levels=design)
fit2 <- contrasts.fit(fit, cont.matrix)
fit2 <- eBayes(fit2, 0.01)
tT <- topTable(fit2, adjust="fdr", sort.by="B", number=250)
tT <- topTable(fit2, adjust="fdr", sort.by="B", number=18000)
sessionInfo()
dim(exprs(gset))
tT <- topTable(fit2, adjust="fdr", sort.by="B", number=18200)
gpl <- annotation(gset)
platf <- getGEO(gpl, AnnotGPL=TRUE)
ncbifd <- data.frame(attr(dataTable(platf), "table"))
# replace original platform annotation
tT <- tT[setdiff(colnames(tT), setdiff(fvarLabels(gset), "ID"))]
tT <- merge(tT, ncbifd, by="ID")
tT <- tT[order(tT$P.Value), ]  # restore correct order
tT <- subset(tT, select=c("ID","adj.P.Val","P.Value","t","B","logFC","Gene.symbol","Gene.ID", "Gene.title"))
write.table(tT, file='DE18200.txt', row.names=T, sep="\t")
save.image('Miles3.RData')
exi<-exprs(gset)
save(sml,tT,exi,file='imports.rda')
savehistory("Geo2R.REDO.R")