DESeq2 dds Creation

Question

[cgp09741]

Hello,

I am trying to generate a dds out of a count matrix file. I haven successfully imported the count matrix for "countData" as well as the metadata for "colData" but when I run "DESeqDataFromMatrix" I receive the error

Error in SummarizedExperiment(assays = SimpleList(counts = countData), : the rownames and colnames of the supplied assay(s) must be NULL or identical to those of the SummarizedExperiment object (or derivative) to construct

Below are the code lines I am running to get to this point, any help would be appreciated to generate a dds so I can run DESeq2.

raw_counts <- read.csv("raw_counts.csv", header = TRUE, row.names = 1)
countdata <- as.matrix(raw_counts)
coldata <- read.csv("metadata.csv", header = T, row.names = 1)
dds <- DESeqDataSetFromMatrix(countData = countdata,
                          colData = coldata,
                          design = ~ sample)

Thank you.

Answer 1

[ATpoint]

object 'countData' not found

It must be countdata, with a lowercase d as in the 2nd line of code.

[cgp09741]

Yes, thank you I caught that right before seeing this! I get a new error when I correct that which is:

Error in SummarizedExperiment(assays = SimpleList(counts = countData), : the rownames and colnames of the supplied assay(s) must be NULL or identical to those of the SummarizedExperiment object (or derivative) to construct

[jared.andrews07]

Check that all(rownames(coldata) %in% colnames(countdata)). If it returns FALSE, your colData rownames don't match the column names for the counts, so the samples can't be properly associated with the sample-specific metadata.

Correcting that should solve the error.

[cgp09741]

Great, thank you. It seems they are not aligned. What code would align the two?

[ATpoint]

In the simplest case you make an intersect, to get the samples that are in both countdata and coldata and then sort them. Something like:

#/ for example (untested)
i <- intersect(colnames(countdata), rownames(coldata))`
countsdata2 <- countdata[,i]
coldata2 <- coldata[i,]