Hello I am using DiffBind to identify consensus binding sites for multiple transcription factors that have biological replicates. In addition, I want to investigate the overlap of binding sites across the transcription factors. I am able to call consensus peaks for each transcription factor and calculate the overlap rate and plot overlaps with dba.plotVenn but I am getting an error from GRanges when trying to extract the actual overlapping peaks using dba.overlap and DBA_OLAP_PEAKS. Any suggestions on why I am getting this error? Thanks Matt > #load datasets > chip= dba(sampleSheet="chip_datasets_testing.csv", peakCaller="bed") a4142.1 a21 ARK2 1 bed a4142.2 a22 ARK2 2 bed a0304.1 a23 ARK2 1 bed a0304.2 a24 ARK2 2 bed r4748.1 r11 REV 1 bed r8586.1 r21 REV 1 bed r8586.2 r22 REV 2 bed c4344.1 c11 PCN 1 bed c4344.2 c12 PCN 2 bed c4546.1 c21 PCN 1 bed c4546.2 c22 PCN 2 bed a3738.1 a11 ARK1 1 bed a3738.2 a12 ARK1 2 bed a3940.2 a14 ARK1 2 bed a3940.1 a13 ARK1 1 bed > > #create consensus peaks and plot overlap > chip = dba.peakset(chip, consensus = DBA_TREATMENT, minOverlap = 0.5) Add consensus: ARK2 Add consensus: REV Add consensus: PCN Add consensus: ARK1 > > chip 19 Samples, 12123 sites in matrix (29678 total): ID Condition Treatment Replicate Peak.caller Intervals 1 a4142.1 a21 ARK2 1 bed 3239 2 a4142.2 a22 ARK2 2 bed 2026 3 a0304.1 a23 ARK2 1 bed 2718 4 a0304.2 a24 ARK2 2 bed 581 5 r4748.1 r11 REV 1 bed 6958 6 r8586.1 r21 REV 1 bed 595 7 r8586.2 r22 REV 2 bed 869 8 c4344.1 c11 PCN 1 bed 8526 9 c4344.2 c12 PCN 2 bed 803 10 c4546.1 c21 PCN 1 bed 5524 11 c4546.2 c22 PCN 2 bed 5320 12 a3738.1 a11 ARK1 1 bed 7443 13 a3738.2 a12 ARK1 2 bed 7004 14 a3940.2 a14 ARK1 2 bed 5697 15 a3940.1 a13 ARK1 1 bed 761 16 ARK2 a21-a22-a23-a24 ARK2 1-2 bed 2030 17 REV r11-r21-r22 REV 1-2 bed 548 18 PCN c11-c12-c21-c22 PCN 1-2 bed 3500 19 ARK1 a11-a12-a14-a13 ARK1 1-2 bed 6210 > > dba.overlap(chip,c(16,17,18,19), mode=DBA_OLAP_RATE) [1] 10006 1733 327 11 > > chip.OL = dba.overlap(chip, c(16,17,18,19),mode=DBA_OLAP_PEAKS) Error in validObject(.Object) : invalid class ???GRanges??? object: NROW(strand(x)) != length(x) -- output of sessionInfo(): > sessionInfo() R version 3.0.2 (2013-09-25) Platform: x86_64-w64-mingw32/x64 (64-bit) locale: [1] LC_COLLATE=English_United States.1252 [2] LC_CTYPE=English_United States.1252 [3] LC_MONETARY=English_United States.1252 [4] LC_NUMERIC=C [5] LC_TIME=English_United States.1252 attached base packages: [1] parallel stats graphics grDevices utils datasets methods [8] base other attached packages: [1] DiffBind_1.8.2 Biobase_2.22.0 GenomicRanges_1.14.3 [4] XVector_0.2.0 IRanges_1.20.5 BiocGenerics_0.8.0 loaded via a namespace (and not attached): [1] amap_0.8-7 bitops_1.0-6 caTools_1.16 [4] edgeR_3.4.0 gdata_2.13.2 gplots_2.12.1 [7] gtools_3.1.1 KernSmooth_2.23-10 limma_3.18.2 [10] RColorBrewer_1.0-5 stats4_3.0.2 tools_3.0.2 [13] zlibbioc_1.8.0 -- Sent via the guest posting facility at bioconductor.org.

Hi Matt- Your code looks good -- this looks like a bug. It must be something about the specific 4-way overlap that you are doing as I can't reproduce it with some datasets I have. Is there a way you can share the DiffBind Object ("chip") with me so I can debug it? Dropbox perhaps? Cheers- Rory On 14/11/2013 01:04, "Matt Zinkgraf [guest]" <guest at="" bioconductor.org=""> wrote: > >Hello >I am using DiffBind to identify consensus binding sites for multiple >transcription factors that have biological replicates. In addition, I >want to investigate the overlap of binding sites across the transcription >factors. I am able to call consensus peaks for each transcription factor >and calculate the overlap rate and plot overlaps with dba.plotVenn but I >am getting an error from GRanges when trying to extract the actual >overlapping peaks using dba.overlap and DBA_OLAP_PEAKS. Any suggestions >on why I am getting this error? > >Thanks >Matt > >> #load datasets >> chip= dba(sampleSheet="chip_datasets_testing.csv", peakCaller="bed") >a4142.1 a21 ARK2 1 bed >a4142.2 a22 ARK2 2 bed >a0304.1 a23 ARK2 1 bed >a0304.2 a24 ARK2 2 bed >r4748.1 r11 REV 1 bed >r8586.1 r21 REV 1 bed >r8586.2 r22 REV 2 bed >c4344.1 c11 PCN 1 bed >c4344.2 c12 PCN 2 bed >c4546.1 c21 PCN 1 bed >c4546.2 c22 PCN 2 bed >a3738.1 a11 ARK1 1 bed >a3738.2 a12 ARK1 2 bed >a3940.2 a14 ARK1 2 bed >a3940.1 a13 ARK1 1 bed >> >> #create consensus peaks and plot overlap >> chip = dba.peakset(chip, consensus = DBA_TREATMENT, minOverlap = 0.5) >Add consensus: ARK2 >Add consensus: REV >Add consensus: PCN >Add consensus: ARK1 >> >> chip >19 Samples, 12123 sites in matrix (29678 total): > ID Condition Treatment Replicate Peak.caller Intervals >1 a4142.1 a21 ARK2 1 bed 3239 >2 a4142.2 a22 ARK2 2 bed 2026 >3 a0304.1 a23 ARK2 1 bed 2718 >4 a0304.2 a24 ARK2 2 bed 581 >5 r4748.1 r11 REV 1 bed 6958 >6 r8586.1 r21 REV 1 bed 595 >7 r8586.2 r22 REV 2 bed 869 >8 c4344.1 c11 PCN 1 bed 8526 >9 c4344.2 c12 PCN 2 bed 803 >10 c4546.1 c21 PCN 1 bed 5524 >11 c4546.2 c22 PCN 2 bed 5320 >12 a3738.1 a11 ARK1 1 bed 7443 >13 a3738.2 a12 ARK1 2 bed 7004 >14 a3940.2 a14 ARK1 2 bed 5697 >15 a3940.1 a13 ARK1 1 bed 761 >16 ARK2 a21-a22-a23-a24 ARK2 1-2 bed 2030 >17 REV r11-r21-r22 REV 1-2 bed 548 >18 PCN c11-c12-c21-c22 PCN 1-2 bed 3500 >19 ARK1 a11-a12-a14-a13 ARK1 1-2 bed 6210 >> >> dba.overlap(chip,c(16,17,18,19), mode=DBA_OLAP_RATE) >[1] 10006 1733 327 11 >> >> chip.OL = dba.overlap(chip, c(16,17,18,19),mode=DBA_OLAP_PEAKS) >Error in validObject(.Object) : > invalid class ???GRanges??? object: NROW(strand(x)) != length(x) > > -- output of sessionInfo(): > >> sessionInfo() >R version 3.0.2 (2013-09-25) >Platform: x86_64-w64-mingw32/x64 (64-bit) > >locale: >[1] LC_COLLATE=English_United States.1252 >[2] LC_CTYPE=English_United States.1252 >[3] LC_MONETARY=English_United States.1252 >[4] LC_NUMERIC=C >[5] LC_TIME=English_United States.1252 > >attached base packages: >[1] parallel stats graphics grDevices utils datasets methods >[8] base > >other attached packages: >[1] DiffBind_1.8.2 Biobase_2.22.0 GenomicRanges_1.14.3 >[4] XVector_0.2.0 IRanges_1.20.5 BiocGenerics_0.8.0 > >loaded via a namespace (and not attached): > [1] amap_0.8-7 bitops_1.0-6 caTools_1.16 > [4] edgeR_3.4.0 gdata_2.13.2 gplots_2.12.1 > [7] gtools_3.1.1 KernSmooth_2.23-10 limma_3.18.2 >[10] RColorBrewer_1.0-5 stats4_3.0.2 tools_3.0.2 >[13] zlibbioc_1.8.0 > >-- >Sent via the guest posting facility at bioconductor.org.