Hi,
I'm trying to use dba.contrast and dba.analyze to find differential binding sites within specific regions (ie  promoter region ranging from 1000bp upstream of TSS and 500 bp downstream of TSS for a set of genes). I realized that dba.count can use pre-defined regions with the peaks argument so I did something like the following:

# create dba object

TF.peaks <- dba(sampleSheet = "experiments.csv",peakCaller = "macs",peakFormat = "bed",scoreCol = 5 );

# create a GRanges object with specific region

genes <- read.delim('gene_list.txt', as.is = TRUE);
grange <- makeGRangesFromDataFrame(genes);

promoter <- promoters(grange, upstream = 1000, downstream = 500);

# count the reads in the pre-defined region before running dba.contrast and dba.analyze

TF.counts <- dba.count(DBA = TF.peaks, peaks = promoter, score = DBA_SCORE_RPKM);

I got no error message, however, my TF.counts chromosome regions are beyond the range of some chromosome sizes. For example:
range(TF.counts$peaks[[1]]$Start[TF.counts$peaks[[1]]$Chr == 'chr22'])
[1] 22082728 102264308
I double checked my input TF.peaks and promoter which all have the correct chr22 size
range(TF.peaks$peaks[[1]]$V2[TF.peaks$peaks[[1]]$V1 == 'chr22'])
[1] 17639813 51222046
promoter.sub <- promoter[promoter@seqnames == 'chr22',]
range(promoter.sub@ranges@start)
[1] 17081777 51221591
Any idea what's causing the issue? Or is there a better way to look at a differential binding site for specific regions? (I'm using R v3.3.1 and DiffBind v2.2.12)
Many thanks,

Sylvia

If you can send me the intervals you are passing in as promoter, I can take a look at what is happening. If possible, it would be good to have a copy of the main DBA object TF.peaks -- if this is too large, you can send me a download link (Dropbox or similar).

Cheers-

Rory

rory.stark @ cruk.cam.ac.uk

P.S. It is best to tag DiffBind questions with the DiffBind tag as well...

I've had a look and identified a bug, which should be fixed by the end of the week in DiffBind 2.6.2.

I see you are using an older version, I can provide a workaround. You just need to pass in the intervals as a data.frame, with the chromosome names (first column) as character strings:

> promoter <- promoters(grange, upstream = 1000, downstream = 500)
> promoterDF <- data.frame(promoter)
> promoterDF[,1] <- as.character(promoterDF[,1])
> TF.counts <- dba.count(DBA = TF.peaks, peaks = promoterDF, score = DBA_SCORE_RPKM)

Cheers-

Rory