Hi wondering how I can use the following DESeq2 logic within the framework of DiffBind for calling differential peaks and looking at multiple interactions:

to look at interaction--
dds<-DESeqDataSetFromMatrix(countData = countData,
                           colData = colData,
                           design = ~ sex + treatment + sex:treatment)
dds$sex<-relevel(dds$sex, ref="female")
dds$age<-relevel(dds$treatment, ref="CO")

dds <- DESeq(dds)
resultsNames(dds)
results(dds, name="sexmale.treatment")

Currently DiffBind only handles fairly simple designs. I'm looking to upgrade this in future, but the original idea was that if you wanted to do more complex modelling, you should use the underlying DE package (DESeq2) directly.

Cheers-

Rory

Well, 3 years later, the latest release of DiffBind finally supports unrestricted models, as well as an clean interface for retrieving DESeq2 analysis objects...

You should be able to use any model design and set up any contrasts that you are able to in DESeq2 directly. You may need to set up some compound factors to get the contrast you want. It can also be helpful to check using bGetCoefficients=TRUE to understand what the coefficients are.

You can relevel the metadata using the reorderMeta parameter to dba.contrast(). You can just set the reference level or all the levels for a metadata factor. See the help page for dba.contrast().

You can specify any contrast that you could using DESeq2 directly. You have to specify them one at a time with separate calls to dba.contrast().

For example:

k4model <- dba.contrast(k4norm,design="~Tissue+Condition+Tissue:Condition", 
                        reorderMeta=list(Tissue="A",Condition="1"))
k4.model.coeffs <- dba.contrast(k4model, bGetCoefficients=TRUE )
k4model <- dba.contrast(k4model, contrast=list("Tissue_C_vs_A","Tissue_B_vs_A" ))
k4model <- dba.contrast(k4model, contrast="TissueB.Condition2") # implies comparison to reference "TissueA.Condition1"

After setting up all the contrasts of interest, you can call dba.analyze() to compute the model and perform the testing for each contrast. Then you can get the results by calling dba.report() or one of the plotting functions. These need to be called in turn for each contrast:

k4model <- dba.analyze(k4model)
k4model.contrast1 <- dba.report(k4model, contrast=1)
k4model.contrast2 <- dba.report(k4model, contrast=2)

Dear R. stark,

Following your answer, these are my steps :

k4dba <- dba(sampleSheet = chipk4)

k4count <- dba.count(k4dba,bParallel = T)

k4norm <- dba.normalize(k4count)

k4model <- dba.contrast(k4norm,design="~Tissue+Condition+Tissue:Condition",bGetCoefficients = T)

The coefficient obtained are:

"Intercept" "Tissue_C_vs_A" "Tissue_B_vs_A" "Condition_3_vs_1" "Condition_2_vs_1" "TissueC.Condition3" "TissueB.Condition3" "TissueC.Condition2" "TissueB.Condition2"

Currently when I want to make a contrast on DESeq2 for example: What is the difference between Tissue C and B ? --> contrast = list( "Tissue_C_vs_A" "Tissue_B_vs_A" ) Or if I want the interaction i.e, the condition 2 on the tissue B --> contrast = c("TissueB.Condition2")

For DiffBind, how to make these contrasts ? Using the dba.contrast() or dba.report() and before the dba.analyse() ?

Moreover, using a simply relevel() can I modify the reference level of the Tissue or Condition ?

Thanks

Dear R. stark,

Thank you, this is perfect for the next steps !

Thank again

P. Marin