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DESeq2 on GEO dataset

Hi, I am looking to perform Differential Gene expression analysis using R and GEO datasets. But I couldn't find the count matrices in the datasets, it is a raw data and need preprocessing. Can anyone help me how to use DESeq2 libary to preprocess the GEO dataset for Differential gene expression analysis.

rna-seq geo r deseq2 preprocessing
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updated by Biostar Biostar • written by hariprasadp.iitkgp • hariprasadp.iitkgp

1 answer

Okay, that is a microarray, so, you cannot use DESeq2. You should use limma.

To retrieve the already-normalised data, you can simply use:

library(Biobase)
library(GEOquery)
gset <- getGEO("GSE3821", GSEMatrix =TRUE, getGPL=FALSE)
if (length(gset) > 1) idx <- grep("GPL90", attr(gset, "names")) else idx <- 1
gset <- gset[[idx]]

To see the metadata:

pData(gset)
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written by Kevin Blighe Kevin Blighe

Nevermind! I realised that I was using microarray, so wouldn't be able to use it either

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written by oludhe • oludhe

Okay, that is a microarray, so, you cannot use DESeq2. You should use limma.

Same applies to your question. DESeq2 is not for microarrays.

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written by ATpoint ATpoint

Hi Kevin,

Just to jump on this thread rather than make a new one, I am having a bit of an issue actually using the eset in DESeq2. I downloaded an expression set, transformed it into a summarizedexperiment object, but getting into a deseq object is giving me errors about the counts. Here is the input code:

 library(Biobase)
library (GEOquery)
library(DESeq2)

gsm <- getGEO('GSE116904') #RNA-seq data

gse <- gsm [[1]]

summ_exp <- makeSummarizedExperimentFromExpressionSet(gse)
ddse <- DESeqDataSet(summ_exp, countData = as.matrix(countData),design = ~ title)

And here is the error:

> ddse <- DESeqDataSet(summ_exp, design = ~ title)
renaming the first element in assays to 'counts'
Error in DESeqDataSet(summ_exp, design = ~title) : 
  some values in assay are not integers

When I specify countData:

ddse <- DESeqDataSet(summ_exp, countData = as.matrix(assay(summ_exp)),design = ~ title)

I get the following error:

Error in DESeqDataSet(summ_exp, countData = as.matrix(assay(summ_exp)),  : 
  unused argument (countData = as.matrix(assay(summ_exp)))

I'm not really sure how to get around this.

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written by oludhe • oludhe

The raw counts are at the bottom of https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE116904.

Use these and feed them into DESeq2 with DESeqDataSetFromMatrix, follow the manual for this. You are still trying to use microarray functions (getGEO) for RNA-seq data. The RNA-seq data are not available by this function. Check the content of gse it has zero rows so no genes = no counts.

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written by ATpoint ATpoint

Yes, you need raw counts. The data obtained via getGEO may be empty or represent the normalised / transformed data.

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written by Kevin Blighe Kevin Blighe

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