Question: Differential Gene Analysis
0
gravatar for kalyanimeha
22 months ago by
kalyanimeha30
New Delhi INDIA
kalyanimeha30 wrote:

I had generated volcano plot by using limma package, but how to get a list of a gene that is upregulated or downregulated separately.

rna-seq chip-seq R • 621 views
ADD COMMENTlink modified 22 months ago by h.mon29k • written 22 months ago by kalyanimeha30
0
gravatar for h.mon
22 months ago by
h.mon29k
Brazil
h.mon29k wrote:

Look at decideTests:

An object of class TestResults. This is essentially a numeric matrix with elements -1, 0 or 1 depending on whether each t-statistic is classified as significantly negative, not significant or significantly positive.

Or look at the logFC column from topTable.

ADD COMMENTlink written 22 months ago by h.mon29k

This is the command I am using to generate volcano plot so now which command should i use to get down and up-regulated gene.

Differential expression analysis with limma

library(Biobase)

library(GEOquery)

library(limma)

load series and platform data from GEO

gset <- getGEO("GSE34747", GSEMatrix =TRUE, AnnotGPL=FALSE)

if (length(gset) > 1) idx <- grep("GPL15069", 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

gsms <- "000111"

sml <- c()

for (i in 1:nchar(gsms)) { sml[i] <- substr(gsms,i,i) }

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

sml <- paste("G", sml, sep="") # set group names

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)

volcanoplot(fit2)

ADD REPLYlink written 22 months ago by kalyanimeha30
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