Bioconductor Forum

Currently I'm running DiffBind using bam/bed files that have been filtered to keep only the highly reproducible peaks (HR-peaks), obtained using...However, in a discussion with my colleagues a point was raised mentioning that the input for DiffBind should be all peaks on the dataset and not only the HR ones, resulting in way less DB peaks for some of our marks/conditions...I find this a little bi…

DiffBind

updated 12 months ago • Oscar

gene in the diseased group is more accessible than the control group? The object is created from the DiffBind package. Thank you so much! ![enter image description here][1] ![enter image description here][2] [1]: /media/images/a55c87a4

DiffBind ATACSeq

updated 12 months ago • Chris

Hi Dr. Stark, I was wondering if I could ask for your assistance with something DiffBind related please? My ChIP-seq experiment consists of five treatment groups (negative control, drug 1, drug 2, drug 3, drug...I do not have access to a high-performance computing cluster so am using the latest version of DiffBind in Galaxy for my analysis. For some reason the Galaxy version of DiffBind (2.10.…

DiffBind

updated 13 months ago • epipally

such that one can look at the plot and see the density along the length of the chromosome. From the diffbind table, what column can I use to plot the read density? Can I use the Fold column? Thank you so much for any help or guidance

Read density DiffBind

updated 13 months ago • bhandary.8590

Hi all, I looked at the GenomicAlignments package but don't know how. Would you have a suggestion? Thank you so much! I don't know why bioconductor automatically switch to follow by message even thought I chose by email.

ATACSeq

updated 13 months ago • Chris

Hi all, Would you suggest how to fix this error? I have only 2 replicates for the control and 2 replicates for the diseased group. Thank you so much. result <- dba.analyze(samples, bParallel=FALSE) [W::hts_idx_load3] The index file is older than the data file: /labs/PI/atacseq_output_03172023/bwa/merged_library/DISEASED_REP2.mLb.clN.sorted.bam.bai Normalize DESeq2 wit…

ATACSeq

updated 13 months ago • Chris

my hair out trying to figure out how to resolve an issue I am having with reading peaksets into Diffbind. The peaks files I am trying to use are .narrowPeak files derived from MACS2. It is my understanding that Diffbind is...binary operator" ``` I have no "scoreCol" column in my sample sheet. I wondered whether maybe Diffbind was having trouble with the .narowPeak files so substituted thes…

DiffBind

updated 13 months ago • os306

Hello, I am have ChIPseq data from histone marks from 2 different condition (mock and treated with 2 biological replicas each) and the aim of my analysis is to study whether a particular histone mark is enriched in one condition versus the other. I am new to bioinformatic and I don't have any statistical training and I am trying to teach myself how to normalise the data and perform the diff…

DESeq2 ChIPSeq DiffBind edgeR HistoneModification

updated 13 months ago • Giulia

Hello all, Seem there is something wrong with my samples samples <- read.csv("samplesheet_DiffBind.csv") result <- dba.analyze(samples) Error in peaks[, pCol]/max(peaks[, pCol]) : non-numeric argument to binary operator I understand what this error means but don't know what is the cause. Would you suggest to me how to fix this? Thank you so much!

DiffBind ATACSeq

updated 13 months ago • Chris

Hello all, I am trying to use DiffBind on my ATAC seq data. However, the code in the manual uses ChIP seq data. Would you advise me on what to do for a learner...Hello all, I am trying to use DiffBind on my ATAC seq data. However, the code in the manual uses ChIP seq data. Would you advise me on what to do for a learner? Thank

ATACSeq

updated 13 months ago • Chris

Hello, I am have ChIPseq data from histone marks from 2 different condition (mock and treated with 2 biological replicas each) and the aim of my analysis is to study whether a particular histone mark is enriched in one condition versus the other. I am new to bioinformatic and I don't have any statistical training and I am trying to teach myself how to normalise the data and perform the diff…

DESeq2 ChIPSeq DiffBind edgeR HistoneModification

updated 13 months ago • Giulia

two groups, and I hope to contrast them to determine differentially methylated regions/genes with DiffBind. The problem lies with the fact that I'm unsure how the data was normalized, if it was at all. Therefore, I would like to

DiffBind

updated 14 months ago • ssankar3

Hi, I want to use DiffBind for differential accessible chromatin regions for my ATACSeq analysis. I am trying to understand the core codes...for this analysis as described in DiffBind manual but getting bit confused due to many interlinked options. If the metadata (information of case, control, replicates...with the command "dba.analyze("File.csv")". With this scenario I have three questions: …

dba.analyze DiffBind @rory-stark-5741

updated 14 months ago • tahir.msajid

differential peak finding, drawing Venn diagram and PCA plot) my m6A-seq data recently using DiffBind. However, as the program is developed originally for analyzing ChIP-seq data, I'm hesitant to apply default settings...events will make aligner generate large gaps between alignments. So maybe some parameters of DiffBind should also be tweaked manually. I'm wondering which parameter should I part…

DiffBind MeRIP-seq m6A-seq

updated 14 months ago • zhiyi

Hi, I'm confused by how DiffBind uses the spike-ins for normalization. My understanding of the manual is that DiffBind calculates the spike-in reads

SpikeIn DiffBind

updated 14 months ago • Weisheng

Hi Rory Stark， DiffBind normalization based on the total reads of a bam file by default, right? And the final results of DiffBind: Conc_Resistant

DiffBind

updated 14 months ago • skyyks123

Hi Rory Stark, I am a little confused about the following questions regarding the use of DiffBind: How does DiffBind handle normalization? Does it use the raw counts from BAM files? Are the total reads counts used...as we analyze RNA-seq read counts, which require raw read counts as input? Is it possible to use DiffBind without spike-in normalization? Thanks in advance

DiffBind

updated 14 months ago • skyyks123

running Diffbind, I don't find any significant differential binding sites. However, if I take the intersection of replicates in each...the other. Those peaks should be statistically significant, but I am not sure I understand why DiffBind did not find any regions that are significant although some regions exist in one group but not the other after taking...the intersection of replicates in eac…

DifferentialPeakCalling ChIPanalyser DiffBind chipseqDB

updated 14 months ago • narzouni

I'm trying to prevent Diffbind from merging overlapping peaks in my bed file (TSS file with exactly 2000 bp widths) by setting DBA$config$mergeOverlap

DiffBind

updated 15 months ago • slrpatty

Is there a way to differentiate between gain peaks and new peaks? I want to be able to get a set of peaks that exist in two sample groups, but higher in one of them and a set of peaks that only appear in one sample but not in the other (new peaks) Thank you very much

ATAC DiffBind ATACSeq

updated 15 months ago • Hadar

I'm comparing knockdown ChIP (3 replicates in the green) versus control ChIP (3 replicates in the orange), but the Fold (in the grey) is much lower than I expected it to be. I know the Fold isn't just Log2FC. Could the low Fold value be caused by shrinkage adjustments due to inconsistencies between the replicates? ![enter image description here][1] [1]: /media/images/4818964b-a81c-497e-8cbb-b7…

DiffBind

updated 15 months ago • slrpatty

I am trying to compare two different conditions (with 2 replicates each) using Diffbind. For my Diffbind output, I'm getting 0 counts for one of the conditions and high counts for the other condition. This...to match my normalized browser tracks. There's clearly a peak around the chromosome position, but Diffbind is calculating it as 0 counts. There's definitely still a change in peak intensity b…

DiffBind

updated 16 months ago • slrpatty

Hello, I am new to DiffBind. I am trying to use dba.count, where ELSDiff is the dba Object after reading the .csv file: ``` ELSDiff <- dba.count(ELSDiff

DiffBind

updated 16 months ago • narzouni

Hi, I'm working with diffbind R package to perform differential analysis on h3k27ac peak data, however, in my case I have to generate a new DBA object...containing custom read count across a number of patient samples. Following diffbind instructions, I tried to generate the DBA object starting from the 'dba' command using a Datasheet with all the required...CD34_control_PE" "CD34_control_PE" …

DiffBind dbaobject

updated 16 months ago • francesco.gandolfi

genome separately. And now I am trying to get normalization factors for each replicate using DiffBind (v. 3.8.3), which I would like to use for differential peak analysis as well as scaling bigwig files. *Qn1:* From what I can...gather, in DiffBind, by including the spikein bam filles in the Spikein column of sampleSheet, I have the option to use the spikein library...spike-in peaks is the corre…

DiffBind SpikeIn Normalization ChIP-seq csaw

updated 17 months ago • spg

Hi, I have generated two sets (two tissues) of histone modification CUT&RUN data, each consists of three replicates of WT and three replicates of a mutant. I successfully made a heatmap using dba.plotProfile and now understand a bit of profileplyr to change some aesthetics. What I couldn't understand were some details. 1. In one tissue (data set 1), the plot shows WT first on …

DiffBind

updated 17 months ago • Junsik

Hi, I'm trying to run DiffBind `3.4.11` on R `4.1.2` on some bed files created using bedtools merge, so they look like: ```r 1 1875658 1934008 EdU_HU_HME1_sgMSL1_R1_peak_54

DiffBind

updated 17 months ago • Pablo

do I need to create something else? Right now this is my R script: ``` library(ChIPQC) library(DiffBind) library(tidyverse) sampleSheet <- read.csv("meta/peaks.xls") db <- dba(sampleSheet =sampleSheet) ``` When I look at the

ChipSeq ChIPSeqData chipseqDB

updated 17 months ago • stacy.genovese

answers to each post are extremely helpful. However, I feel confused to some results while running DiffBind. To better illustrate my problems, I would like to invite you to go through my case. library("DiffBind") samples=read.csv

DiffBind

updated 18 months ago • littlefishes20

Performing `dba.count()` seems to mess up `options('mc.cores')` in the global environment ```r options('mc.cores' = 6L) peaks <- dba(sampleSheet = samples_df) peaks <- dba.count(peaks) class(peaks) # [1] "DBA" options('mc.cores') # $mc.cores # $mc.cores$mc.cores # $mc.cores$mc.cores$mc.cores # [1] 6 ``` This then leads to downstream issues with `dba.blacklist()` and I end u…

DiffBind

updated 18 months ago • Stevie Pederson

Hi, I did an analysis using DiffBind in June 2022 with no errors. I restarted the project last week after updating R and DiffBind and now the normalization...I mentioned above, with an earlier R version (R version 4.1.2 (2021-11-01) -- "Bird Hippie") and DiffBind version (not sure which one) on exactly the same BAM files with the same code I did not have this problem. However, I would

DiffBind Blacklist

updated 18 months ago • Romain

sequence coverage difference between replicates and among samples? I am using DESeq2 and edgeR in DiffBind to compare ATAC-seq samples with different sequencing coverage among samples, from 77 to 25 million reads, and up

DESeq2 edgeR DiffBind

updated 18 months ago • cesar.arenasmena

Hi all (and hopefully Rory would see this. Thanks for the great program) I have ChIP-seq data of 3 replicates of wild type and 3 replicates of my mutant. I'm looking for loci where my protein binds in WT samples (so WT has peaks in the loci) but the mutant has no peaks (the protein doesn't bind to the same loci in the mutants). I think this is what DiffBind manual explained as Occupan…

DiffBind

updated 18 months ago • Junsik

Dear all, and Rory, I would very much appreciate if you could please advise how I shall start the DiffBind workflows from a file with peak counts (binding matrix) as an input into DiffBind. The described workflow in the vignette

DESeq2 edgeR DiffBind

updated 18 months ago • Bogdan

Hello, Thanks for your effort in producing such a good tool for analysis. Recently, I am using Diffbind to characterize differential open chromosome regions with ATAC-seq. I have more than 50 samples, equipping with...Hello, Thanks for your effort in producing such a good tool for analysis. Recently, I am using Diffbind to characterize differential open chromosome regions with ATAC-seq. …

DiffBind

updated 19 months ago • littlefishes20

Hello: Dose anyone have experiences in using DiffBind for spike-in normalization? I have dm6 spike-in for both input and IP samples. I have aligned bam files for each sample...Hello: Dose anyone have experiences in using DiffBind for spike-in normalization? I have dm6 spike-in for both input and IP samples. I have aligned bam files for each sample with both human and drosophila genomes, i.e, …

DiffBind

updated 19 months ago • yuhuihui2011

that we can use for the analysis of differential binding ? (I do not have access to the packages DiffBind or derfinder) ; much thanks, Bogdan

DESeq2 edgeR bigwig limma

updated 19 months ago • Bogdan

Hello! TIA for helping =) so via DiffBind, I import a SampleSheet: ```SS <- DiffBind::dba(sampleSheet = csvFile, scoreCol = 5)``` The sample sheet has 6 bed files, 3 replicates...of one sample and 3 replicates of a second sample. I made the venn diagram: ```Venn <- DiffBind::dba.plotVenn(SS, 1:3)``` everything works great and I get the number of peaks that overlap between th…

chipseq DiffBind

updated 19 months ago • Nicole

K27ac, and K4me1. We carried out peak calling using SEACR and then differential peak calling using Diffbind. Now, we want to go back to the raw peak data and just plot the normalized peak data. Can i use the dba.normalize function...to ChipSeeker so I can plot it. I don't know if this is a naive question because I went through the diffbind tutorial and couldn't find anything specific to normalizi…

Normalization Peakdata DiffBind Cut&Tag

updated 19 months ago • bhandary.8590

Hi, I am trying to look at differential binding of a number of chromatin regulatory proteins via CutandRun only at a **predefined** set of peaks (Approx. 5000 peaks). When I carry out the analysis for different proteins at the same predefined peaks I get a different number of consensus peaks which is always less than 5000. I assumed the analysis would be carried out with the entire predefined…

DiffBind

updated 19 months ago • doherta6

I'm getting an error repeatable error when I try to run code that worked with previous versions of DiffBind (3.4.11) ```r meta <- read_csv("../01b_ChIPQC/HCA_chipqc_meta_final.csv")[c(3:8),] config <- data.frame(minQCth=30, fragmentSize...Traceback returns the dba() function call. This issue is resolved when I revert to R 4.1.2 and DiffBind 3.4.11. Has anyone else encountered this …

DiffBind ChIPQC

updated 20 months ago • ryleehackley

when i put import 12 bed files in one samplesheet, i get 42967 sites in matrix. however, when i remove one of the bed files, the number of sites in matrix changes. the 11 and 12 bed files contain similar peak locations, as some should overlap each other...so I'm not sure if this is a problem because the files contain similar peak locations? any ideas are appreciated! TIA ```NewSampleSheet_DBA …

dba DiffBind

updated 21 months ago • Nicole

with 12 bed files, i get this from str(dba): ```$ totalMerged: int 42967 $ called : num [1:42967, 1:12] 0 1 1 1 0 0 1 1 1 1 ... $ merged : num [1:42967, 1:3] 1 1 1 1 1 1 1 1 1 1 ... ``` with 11 bed files, i get this from str(dba): ```$ totalMerged: int 42967 $ called : num [1:3477, 1:11] 0 0 0 0 0 0 0 0 0 0 ... $ merged : num [1:42967, 1:3] 1 1 1 1 1 1 1 1 1 1 ... ``` wha…

diffbind dba

updated 21 months ago • Nicole

Hello! I am performing a differential analysis on ATAC-seq data in DiffBind. I would like to know if it is possible to plot ellipses around a PCA plot. I generated my plots with the following code

PrincipalComponent DiffBind

updated 22 months ago • Lisa

Hello, I am currently running an analysis using DiffBind on Cut&Run data to identify differentially expressed sites. On several occasions, after running DiffBind with...any significantly differentially bound sites? Below is the output of my command session running DiffBind: ``` > library(DiffBind) LTxding required package: GenomicRanges LTxding required package: stats4 …

Cut&Run DiffBind

updated 22 months ago • gannoncottone

New to R, but enthusiastic to start easing into the world of bioinformatics analysis. Currently partway through my PhD, creating and testing new protocols (ChIP-Seq, bioinformatics analyses) solo in a small research group without an expert or experienced Post-Doc/PI to provide guidance. I've read through the DiffBind Vignette a few dozen times, and followed through a video tutorial hosted on t…

DiffBind

updated 22 months ago • niimabear

Hello! I am currently using DiffBind 3.6.1 and encountering the following error when attempting to analyze my data: ```r > dba <- dba.contrast(dba, design

DiffBind

updated 22 months ago • millerh1

Hi, I am new to diffbind and have a question about the GRange report generated from dba.report() function. in the demonstrated report (tamoxifen.DB

diffbind

updated 22 months ago • jonathanho1989

have ATAC sequencing data for two types of cells, each with three replicates. Get strange results by DiffBind analysis. Why is my volcano map only halfway？ I don't know where I went wrong. Is there a problem with the original data

ATACSeq DiffBind

updated 22 months ago • young

Hi all, I am new to Diffbind and following the workflow from the handbook: https://bioconductor.org/packages/devel/bioc/vignettes/DiffBind...samples have 0 counts for all genes. check the counting script. what is it that went wrong ? my Diffbind version is (DiffBind_3.4.11) Thanks

peakfiles diffbind HMMRATAC

updated 23 months ago • jonathanho1989

Hi Rory, I am using DiffBind to identify sites differentially bound (affinity analysis) by a transcription factor in control (WT), heterozygous...used both default normalization method and spike in normalization. The results are similar for both. DiffBind identifies a good number of sites that are significantly differentially bound between WT and KO. Comparing HET

DiffBind

updated 23 months ago • ipsc-jl

Dear friends, I have been using diffbind to get RLE/TMM normalized counts. I have noticed that the RLE/TMM normalization is performed based on reads subtracted...I have tried following by setting `bSubControl = F`, but it does not work. ```r library(DiffBind) library(tidyverse) h3k27ac_samples <- read.csv("h3k27ac.csv") #use sorted bam h3k27ac <- dba(sampleSheet=h3k27ac_samples

DiffBind

updated 23 months ago • ksong

Hello, I'm running a differential expression profile for a protein using diffbind. I'm curious about one thing, it's not really a problem per se, but the documentation is unclear about a specific part...and the analyze it under the same parameters, I get a different set of results: ```r library(DiffBind) library(tidyverse) library(rtracklayer) library(dplyr) library(stringr) library(BiocParalle…

DifferentialExpression DiffBind

updated 23 months ago • Luca

DiffBind is very simple, see below, and worked flawlessly so far. We however recently updated our Singularity image, and we now...work with a newer DiffBind package version. Unfortunately, the new(er) versions either introduce a new bug (see below) or at least an undocumented...results in 0 consensus peak sets. I have reproducible code that runs just fine when running with DIffBind 2.14, but resu…

DiffBind

updated 24 months ago • chrarnold

not exist, also good. ``` Result: Nope, no chr21 in any of the 8 peak files ## What does DiffBind see? ```r > sample = dba(sampleSheet=read.csv('sample_table.csv'), \ peakFormat='bed', scoreCol=5, bLowerScoreBetter=FALSE

DiffBind

updated 24 months ago • Mehmet

Hi there, I am currently using DiffBind 3.2.7 for an analysis on RNA Polymerase III (RNAPIII) occupancy in the human genome. Because of the biology of RNAPIII

ChIPSeq DiffBind

updated 2.0 years ago • Drew

I am trying out different DiffBind normalization methods for my ATAC-seq dataset in order to select the most suitable one. However, it looks like no...I am trying out different DiffBind normalization methods for my ATAC-seq dataset in order to select the most suitable one. However, it looks like no matter

ATACSeq Normalization DiffBind

updated 2.1 years ago • bers.anthrope

I am trying to analyse a ChIP-Seq data set with `DiffBind`. These are the contents of my sample sheet ("diffbind_samples.csv"): ``` SampleID,Tissue,Factor,Condition,Treatment,Replicate...I am trying to analyse a ChIP-Seq data set with `DiffBind`. These are the contents of my sample sheet ("diffbind_samples.csv"): ``` SampleID,Tissue,Factor,Condition,Treatment

DiffBind

updated 2.1 years ago • nw416

Suppose that I have three conditions to compare in DiffBind, with two replicates per condition. Suppose that the antibody is the same, but one is WT and the other two - mutations

DiffBind

updated 2.1 years ago • Sam

Hi folks, I am new to ChIP-seq analysis hence this may be a basic question. I want to use DiffBind to get peak summits, but after running `dba.count` with my own peaks, I find the number of peaks has decreased. Is this...Hi folks, I am new to ChIP-seq analysis hence this may be a basic question. I want to use DiffBind to get peak summits, but after running `dba.count` with my own peaks, I f…

DiffBind

updated 2.1 years ago • soyabean