Script for analyzing idat (illumina) microarray files with limma, any suggestion?

Question

[Pol]

I have some idat files to analyze, since I haven’t found a complete script for doing it I have wrote one by myself:

library(limma)

# Files and experimental conditions
targets <- readTargets("targets.txt")  # I have two conditions "KO" and "WT"

# Reading data
idatfiles = dir(path="C:/Data", pattern = ".idat")
bgxfile = dir(path = "C:/Data", pattern = ".bgx")
data = read.idat(idatfiles, bgxfile)

# Normalization and background adjustment
data2 <- neqc(data)

# Build the design matrix for the linear modelling function. 
f <- factor(targets$Condition, levels = unique(targets$Condition))
design <- model.matrix(~0 + f)
colnames(design) <- levels(f)

# Apply the intensity values to lmFit. 
fit <- lmFit(data2, design)

# Create a contrast matrix. In this example, all combinations of contrasts can be set up as below. 
contrast.matrix <- makeContrasts("KO-WT",  levels=design)

# Apply this contrast matrix to the modeled data and compute statistics for the data.
fit2 <- contrasts.fit(fit, contrast.matrix)
fit2 <- eBayes(fit2)

# Output the statistics for the dataset and write them to disk for further analysis.
output <- topTable(fit2, adjust="BH", coef="KO-WT", genelist=data2$genes, number=Inf)
write.table(output, file="Results.txt", sep="\t", quote=FALSE)

I have two questions, is it correct? Do you have any suggestion to improve it?

Thank you very much.

Answer 1

[poisonAlien]

Hi, I have written a small automated script for analysis of idat files from Illumina beadarrays (HT12V4 platform, but you can change the platform to yours). You can find it here.

[Pol]

Thank you, I' ll check it

[Ömer An]

Can you please update the link? Thanks.

[poisonAlien]

Try Here

[Ömer An]

I tried but gave up since the debugging takes longer than creating my own script. Some errors below:

topTable = beadAnalyze(idats = c("9533701097_A_Grn.idat","9533701097_B_Grn.idat","9533701097_C_Grn.idat","9533701097_D_Grn.idat"),names = c("H7_2a_2v_A","H7_2a_2v_B","H7_2v_C","H7_2v_D"),condition = c("2a_2v","2a_2v","2v","2v"),ref.condition = "2v", fdr = 0.05, plotPCA = T) Annotating control probes using package illuminaHumanv4.db Version:1.26.0 Show Traceback Rerun with Debug Error in (function (od, vd) : object and replacement value dimnames differ

Error in validObject(.Object) : invalid class “ExpressionSet” object: 1: sampleNames differ between assayData and phenoData invalid class “ExpressionSet” object: 2: sampleNames differ between phenoData and protocolData

Answer 2

[Gordon Smyth]

Your read.idat call couldn't work because you specified a path to dir but not to read.idat.

read.idat and neqc are the only function calls specific to Illumina arrays. Once you run them, it's just like any other limma analysis.