error while plot data with deSeq2

Question

[bioinformaticssrm2011]

Hi,

I am getting an error while plotting figures after running DESeq2. Here is what I have done:

dds <- DESeqDataSetFromMatrix(countData = mydata, colData = meta, design = ~ Diet)
dds <- estimateSizeFactors(dds)
dds <- DESeq(dds)
plotCounts(dds, gene="ENSSSAG00000003723", intgroup=c("Diet") )

Error in .local(object, ...) : unused argument (intgroup = "Diet")

rld <- rlog(dds, blind=TRUE)
plotPCA(rld, intgroup=c("Diet")) +theme_bw()

Error in .local(object, ...) : unused argument (intgroup = "Diet")

But if I remove intgroup from the code, it works, but I need to look at plot with "Diet" groups

> dds
class: DESeqDataSet 
dim: 42350 48 
metadata(1): version
assays(6): counts mu ... replaceCounts replaceCooks
rownames(42350): ENSSSAG00000004313 ENSSSAG00000047403 ... ENSSSAG00000092601 ENSSSAG00000091009
rowData names(31): baseMean baseVar ... maxCooks replace
colnames(48): T3_mc1_tk2_Lh005 T3_mc1_tk3_Lh001 ... T3_mn3_tk2_Lh001 T3_mn3_tk2_Lh007
colData names(8): id Time ... sizeFactor replaceable

[Basti]

Try DESeq2::plotPCA(rld, intgroup=c("Diet")) +theme_bw(), plotPCA should refer in your case to another package

[bioinformaticssrm2011]

Still getting the same error-

DESeq2::plotPCA(rld, intgroup="Diet") +theme_bw()
Error in .local(object, ...) : unused argument (intgroup = "Diet")

But for other plot, it worked when I used this, that's weird...

DESeq2::plotCounts(dds, gene="ENSSSAG00000003723", intgroup="Diet")

[Basti]

Hum strange, and if you set plotPCA <- DESeq2::plotPCA before running plotPCA(rld, intgroup=c("Diet")), do you still have the same error ?

[bioinformaticssrm2011]

yes, I am still getting the same error

[swbarnes2]

Can you just confirm that Diet is indeed one of your ColData headers in dds? We can't see that from what you've shown.

[bioinformaticssrm2011]

yes, its there, as you can see below:

    colData(rld)
    DataFrame with 48 rows and 7 columns
                                   id        Time     Diet Tank_number Sample_Type Sample_Number sizeFactor
                          <character> <character> <factor> <character> <character>     <integer>  <numeric>
    T3_mc1_tk2_Lh005 T3_ctr_tk1_Lh001          T3      ctr         tk1       s1             1   0.639395
    T3_mc1_tk3_Lh001 T3_ctr_tk1_Lh003          T3      ctr         tk1       s1             3   1.163646

[swbarnes2]

what happens if instead of using "Diet" as as string in the command, you use colnames(colData(dds))[3]? Maybe there's a hidden white space in there?

[bioinformaticssrm2011]

Still the same error. If I use any other group for plotting, it still gives the same error.

[swbarnes2]

You might want to ask on the bioconductor support group. Michael Love does read and respond there.