bug in RNAseqDegs_DESeq function

[yeungj234]

Hello, I have encountered a bug when using DESeq instead of limma for differential gene analysis step.

Here is the error message with batch information included:

Here is the error message without batch information included:

If I use the same exact code and phenoData information but with RNAseqDegs_limma, the function works fine and I don't encounter this issue.

I tried to fix the issue by looking at your source code for RNAseqDegs_DESeq and changing it to this:

RNAseqDegs_DESeq = function(counts, phenodata, complist, qval = 0.05) { celltype = phenodata$celltype compList <- complist DEresult = vector(mode = "list", length = length(compList) + 1) names(DEresult) = c(compList, "Overall")

dds <- DESeqDataSetFromMatrix(countData = counts, colData =phenoData@data , design = ~ celltype) 
dds <- estimateSizeFactors(dds)
e <- counts(dds, normalized = TRUE)

for (comp in compList) {
  cs = strsplit(comp, split = "-")[[1]]
  ids = which(celltype %in% cs)
  tmpcounts = counts[, ids]
  tmpph = as(phenodata[ids, ], "data.frame")

  if (is.null(phenodata$batch) | length(unique(phenodata$batch)) == 1){
    dds = DESeqDataSetFromMatrix(countData = data.frame(tmpcounts), colData = tmpph, design = ~ celltype)
  }else{
    dds = DESeqDataSetFromMatrix(countData = data.frame(tmpcounts), colData = tmpph, design = ~ batch + celltype)
  }
  dds = DESeq(dds)
  DEtable = results(dds, alpha = qval)
  DEtable =  data.frame(DEtable[complete.cases(DEtable), ])
  DEtable = DEtable[order(DEtable$pvalue), ]
  rank_vector = abs(DEtable$stat); names(rank_vector) = rownames(DEtable)
  degs = rownames(DEtable[DEtable$padj < qval, ])
  tmpDErslt <- list(table = DEtable, rank_vector = rank_vector, degs = degs)
  DEresult[[comp]] = tmpDErslt
}
DEresult$Overall = list(table = DEtable, rank_vector = rank_vector, degs = degs, e = e)
return(DEresult) }

This code then allows for DESeq to run properly if only celltype information is provided in phenoData but not batch information. I'm not sure how to fix the problem at this point but since my other analyses have been using DESeq and not limma, I would like to use NetAct with DESeq preferably.

[yeungj234]

note: doing DESEq with the code I provided doesn't allow you to proceed further with downstream steps since I get this error when trying to run the TF_Activity function(). Please advise.

[iamcorrinne]

I also receive the same error message when trying to use RNAseqDegs_DESeq, and I am interested in a fix for this.

[iamcorrinne]

@vivekkohar @lusystemsbio any insight here?

[YukaiYou]

note: doing DESEq with the code I provided doesn't allow you to proceed further with downstream steps since I get this error when trying to run the TF_Activity function(). Please advise.

We have updated this function, and it should work normally after reinstallation. By the way, I think you should use cell type without batch

[YukaiYou]

@vivekkohar @lusystemsbio any insight here?

I also receive the same error message when trying to use RNAseqDegs_DESeq, and I am interested in a fix for this.

After reinstalling, the function should work properly since we have implemented updates.