Hi everyone. I'm new at Diffbind. I'm trying to analyze my data but I noticed that if I try to run the analysis more times on the same data, the results change. How is it possible? I clean the environment everytime because initially I thought that it could be a cache memory problem. Any hint?

This is the code

samplesdbprova<-read.csv("gruppisenza1421.csv")
dbObjprova <- dba(sampleSheet=samplesdbprova)
dbObjprova <- dba.count(dbObjprova,bUseSummarizeOverlaps=TRUE, minOverlap=2)
contrastprova <- dba.contrast(dbObjprova, dbObjprova$masks$SIRT630W, dbObjprova$masks$W30,"SIRT630w", "wt30")
bObjprova <- dba.analyze(contrastprova, method=DBA_ALL_METHODS)

In this case I uploaded a matrix which has 18 samples divided in 4 conditions. For the diff analysis I selected just two of the four conditions (the analysis was carried out on 6 samples vs 5 samples).

I tried also to prepare a matrix for each analysis but the problems still are present.

The coloumns of the object are these one

SampleID,Factor,Replicate,Condition,bamReads,Peaks, PeakCaller, PeakFormat

This is the information about my Rstudio version

R version 3.6.1 (2019-07-05)
Platform: x86_64-w64-mingw32/x64 (64-bit)
Running under: Windows 10 x64 (build 17763)
Matrix products: default
locale:
[1] LC_COLLATE=Italian_Italy.1252  LC_CTYPE=Italian_Italy.1252    LC_MONETARY=Italian_Italy.1252 LC_NUMERIC=C                  
[5] LC_TIME=Italian_Italy.1252    
attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     
loaded via a namespace (and not attached):
[1] compiler_3.6.1 tools_3.6.1   

Thanks Francesca

What exactly is different about the results? How are you comparing? How different are they?

To narrow down what might be happening, first we can identify if the change is happening in the counting or the analysis phases.

You can run the last line multiple times:

bObjprova <- dba.analyze(contrastprova, method=DBA_ALL_METHODS)

and see if you get the same results each time (you could write some code to compare the most recent result with the previous one). If you get different results, it is in the analysis phase and I can take a look at that if you send me a link to the DBA object.

If you keep getting the same results, then it should be in the counting phase. You can run the last three commands multiple times:

dbObjprova <- dba.count(dbObjprova,bUseSummarizeOverlaps=TRUE, minOverlap=2)
contrastprova <- dba.contrast(dbObjprova, dbObjprova$masks$SIRT630W, dbObjprova$masks$W30,"SIRT630w", "wt30")
bObjprova <- dba.analyze(contrastprova, method=DBA_ALL_METHODS)

when you have two DBA objects with different results, you can send them to me and I can try to see how they are different.

Cheers- Rory