Question: DiffBind without replicates
0
gravatar for zhangdengwei
27 days ago by
zhangdengwei0 wrote:

Hi,

I am finding the differential peaks using DiffBind, but my sample has no replicates. And my SampleSheet is

> dbObj <- dba(sampleSheet="SampleSheet.csv")
trisomy_21 fibroblasts trisomy_21 trisomy_21 trisomy_21 1 narrow
euploid fibroblasts euploid euploid euploid 1 narrow
> dbObj
2 Samples, 33153 sites in matrix (47495 total):
          ID      Tissue     Factor  Condition  Treatment Replicate Caller Intervals
1 trisomy_21 fibroblasts trisomy_21 trisomy_21 trisomy_21         1 narrow     40820
2    euploid fibroblasts    euploid    euploid    euploid         1 narrow     44391

When performing the differential analysis, I got the following failure

> dbObj <- dba.contrast(dbObj, categories=DBA_FACTOR,minMembers = 1)
Error in dba.contrast(dbObj, categories = DBA_FACTOR, minMembers = 1) : 
  minMembers must be at least 2. Use of replicates strongly advised.
> dbObj <- dba.contrast(dbObj, categories=DBA_FACTOR,minMembers = 2)
Warning message:
No contrasts added. Perhaps try more categories, or lower value for minMembers. 
> dbObj <- dba.analyze(dbObj)
Error in pv.DBA(DBA, method, bSubControl, bFullLibrarySize, bTagwise = bTagwise,  : 
  Unable to perform analysis: no contrasts specified.
In addition: Warning message:
No contrasts added. Perhaps try more categories, or lower value for minMembers. 
> dbObj <- dba.contrast(dbObj, categories=DBA_CONDITION)
Warning message:
No contrasts added. Perhaps try more categories, or lower value for minMembers. 
> dbObj <- dba.contrast(dbObj, categories=DBA_CONDITION, minMembers = 1)
Error in dba.contrast(dbObj, categories = DBA_CONDITION, minMembers = 1) : 
  minMembers must be at least 2. Use of replicates strongly advised.

So, replication is requisite for DiffBind? If not, what can I do for this case? Thanks very much.

diffbind • 51 views
ADD COMMENTlink modified 27 days ago by Rory Stark2.9k • written 27 days ago by zhangdengwei0
Answer: DiffBind without replicates
1
gravatar for Rory Stark
27 days ago by
Rory Stark2.9k
CRUK, Cambridge, UK
Rory Stark2.9k wrote:

Yes, replicates are required to do any kind of statistical analysis. Replicates are required to estimate the variance in the data and calculate confidence statistics such as p-values/FDRs.

Without replicates, you can do some exploratory analysis of overlapping peaks (occupancy analysis). For example using dba.plotVenn(). But not knowing if your data represents an outlier, combined with the inherent noisiness of peak calling, means you will have to have another way to validate any "differential" peaks you identify.

ADD COMMENTlink written 27 days ago by Rory Stark2.9k

Many thanks for your detailed interpretation. I got it.

ADD REPLYlink written 26 days ago by zhangdengwei0
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