I am trying to analyze the differential peaks in my ATAC-seq data. I have ATAC-seq analysis on 2 biological conditions with 3 replicates in each condition. Below is my sample info:
```
ID Factor Condition Replicate Intervals
1 ATACctrl1 ctrl1 ctrl 1 8654
2 ATACctrl2 ctrl2 ctrl 2 6750
3 ATACctrl3 ctrl3 ctrl 3 5962
4 ATACe…
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When running dba.normalize in diffbind I am receiving the follow error:
```if (!requireNamespace("BiocManager", quietly = TRUE))```
```install.packages("BiocManager...BiocManager::install("DiffBind")```
```library(DiffBind)```
# Load sample sheet
```sampleSheet <- read.csv("/Users/vwimalasena/Documents/Ewald/ChIP-seq/DiffBind...Create a DBA object
```dbaObj <- dba(sampleSheet="/…
updated 12 days ago • Virangika
respective normalisation factors exactly, or the inverse of the normalization factors generated from diffBind?
Thank you!
================
Codes:
```r
library(DiffBind)
library(edgeR)
library(tidyverse)
library(dplyr)
samples <- read.csv('Sample.csv
updated 15 days ago • Henry
function, and subsequently dba.analyze(). I basically followed the steps on page 45 of the Diffbind documentation. https://bioconductor.org/packages/release/bioc/vignettes/DiffBind/inst/doc/DiffBind.pdf
However...or scripts of runnning successful offset and loess normalization?
Thank you!
```r
library(DiffBind)
library(edgeR)
library(tidyverse)
library(dplyr)
samples <…
updated 16 days ago • Henry
Hi,
I am trying to understand PCA plot I generated using Diffbind.
I used this code
ss <- read.csv("samplesheet.csv")
obj <- dba(sampleSheet=ss, scoreCol=5, minOverlap=1)
# get counts (mapQCth...DESeq2 and plotting PCA on both vst transformed counts and raw counts. The PCA obtained from diffbind looks more similar to raw counts than vst.
So what I am doing wr…
Hi all,
I'm new to **diffbind**.
I've been given a set of **ATAC-seq** data computed with [nf-core/atacseq][1] and asked if there is any peak associated with
Hi,
I have been working with Diffbind to identify differential binding of H3K9me3 due to ethanol treatment. We are using both males and females, and initially
updated 5 weeks ago • erbrocato
I am retrieving the normalized counts of reads in peaks (scores) from DiffBind, as explained in [this][1] Bioconductor post :
```
dba.peakset(dbObj_norm, bRetrieve=TRUE ) %>% as.data.frame() %>% head()
seqnames
updated 6 weeks ago • Sam
set num cores and how many reads to store in memory
diffbind$config$cores <- db_config$cores-1
diffbind$config$yieldSize <- 20000000
# get counts
diffbind$config$scanbamparam...FALSE, hasUnmappedMate=FALSE, isMinusStrand=NA, isMateMinusStrand=NA))
# get counts
diffbind <- dba.count(diffbind, summits=DB_summits, bUseSummarizeOverlaps = TRUE, bParallel = TRUE)
pr…
updated 6 weeks ago • red_bricks
Hi!
I am trying to run Diffbind with DESeq2 on my Cut&Run dataset. I have tried multiple normalization methods in dba.count() and dba.normalize
updated 8 weeks ago • be308df4
running the same code without peaks=NULL, score=DBA_SCORE_READS
I am using R version 4.3.1 and DiffBind 3.10.1.
Help is very much appreciated
I'm pretty new to ATAC-Seq analysis so I'm sorry if my question is dumb.
I did read through the DiffBind vignette and I see that Deseq2 was said to be more preferred. When I tried running my data using both EdgeR and Deseq2
Dear all, I am using DiffBind for ChIP-Seq and CUTnTag analysis and would like to switch off subtracting reads of control samples. For this, I set...bSubControl=FALSE` in `dba.count()`. However, DiffBind results are exactly identical no matter if I set bSubControl to TRUE or FALSE. I tried this for several R and DiffBind
updated 3 months ago • frueher
Hi,
I am running Diffbind differential analysis of ATAC-seq samples.
I am testing out the different options in diffbind and I could successfully...KO3_TCDF_peaks.narrowPeak,narrow
My code is
```
# mamba install bioconda::bioconductor-diffbind
library(DiffBind)
SAMPLESHEET="samplesheet_PE_q0.05.csv"
# Minimum overlap for the samples to build the consensus
updated 3 months ago • Aswathy
I'm using diffbind to identify differential binding of CUT&tag peaks of H3K27ac. I obtained scaling factors from ChIPSeqSpikeIn...package. I'm very new to bioinformatic analysis. I know the Diffbind has dba.normalize function which you can specify a numeric normalization factor. Could anyone suggest the code...to specify number for normalization method in dba.normalize using DiffBind? Tha…
updated 3 months ago • Hai
Hi All,
I`m using diffBind to analyze my ATACseq results. I have 2 groups (before and after treatment) in triplicates.
How can I create a plot of...per sample.
Currently I used soGGi package (attached), but I wish to use the data from the diffBind dba.obj and compare it to soGGi.
Thank you !
nucFree <- regionPlot(bamFile = files[[3]], testRanges = TSSs, style = "point",
…
When I set the contrast in DiffBind as
```
# Contrast from normalized object
dbObj_contrast_manual <- dba.contrast(dbObj_norm, design="~Condition")
dbObj_contrast_cells
updated 4 months ago • Sam
I want to run an analysis with minOverlap=1 (all peaks to be included in the consensus peakset). However, dba.blacklist changes the consensus peakset to be based on minOverlap=2. How to run dba.blacklist without changing minOverlap?
```r
dbObj_blacklist <- dba.blacklist(dbObj_initial, blacklist = blacklist_granges, greylist = FALSE)
dbObj_initial$minOverlap
dbObj_blacklist$minOverlap
sessio…
updated 4 months ago • Sam
Hello,
I'm trying to run DiffBind on my CUT&RUN data to get the differentially bound peaks between two conditions (DMSO and inhibitor). In sample_sheet.csv
composition differences during normalization?**
I have read the relevant sections of both the DiffBind and csaw documentation thoroughly, but both assume identical spike-in levels across all samples. Are my thoughts
I am using Diffbind for an ATAC-Seq analysis. My peak caller is MACS2, and here is my sample sheet:**
![my sample sheet][1]
**I run Diffbind with the...idea about this? Thank you so much!**
```
library(BiocParallel)
register(SerialParam())
library(DiffBind)
data <- dba(sampleSheet="sampleSheet.csv")
data <- dba.blacklist(data, blacklist=DBA_BLACKLIST_GRCH38, greylist
Hi,
DiffBind (v3.4.11) dba.analyze function doesn't have bSubControl option anymore?
dba.analyze function (bGreylist=FALSE
updated 6 months ago • r.murakami
Hi,
I ran DiffBind dba.blacklist function to get greylist. Then I noticed that DiffBind returns different grey lists every time I...If there is a hint to solve this problem, please let me know.
Thank you.
```
>library("DiffBind")
>set.seed(42)
>library(BiocParallel)
>register(SerialParam())
>ATAC = dba(sampleSheet="path_to_samplesheet
updated 6 months ago • r.murakami
DiffBind - how to prep the sample matrix when multiple bam files are used by MACS2's -t and -c param
Hi everyone!
When we prepare the sampleSheet matrix for the DiffBind::dba function, we may have 2 columns for bam files - 1 for ChIP bam and 1 for Input/IgG bam.
Here I have 2 bam files for ChIP
Hi all,
I have a questions about bulk ATAc seq that could not find out myself and also on Biostar. I use Diffbind to get the table below (differential accessibility regions) and then use ChipPeakAnno to annotate the peaks in the...I have a questions about bulk ATAc seq that could not find out myself and also on Biostar. I use Diffbind to get the table below (differential accessibility regions) a…
updated 7 months ago • Chris
samples)
dbObj <- dba.count(dbObj, bUseSummarizeOverlaps=TRUE)
```
I use the hg38 genome for diffbind is OK, while wheat genome is so big, how can use diffbind to analyze the wheat project
updated 7 months ago • zgyu12
Hi I am getting this error message after running dba.count()
```r
myDBA <- dba(sampleSheet = subset_data)
myDBA <- dba.count(myDBA)
76 neurons CHD4 Ngn2 Ngn2 1 bed
Computing summits...
Re-centering peaks...
Error in colnames(result)[4:ncol(result)] <- colnames(classes) :
replacement has length zero
In addition: Warning message:
Samples have been merged or lost du…
updated 8 months ago • Mark
Hello I hope you are doing well; I am running into this issue when I am trying to run dba.count().
This is the code I am using:
```r
#Create a new DBA object using the subset data
myDBA <- dba(sampleSheet = subset_data)
myDBA <- dba.count(myDBA, bUseSummarizeOverlaps=FALSE)
print(myDBA)
```
It does the following: Computing summits... Re-centering peaks... Reads will be count…
updated 8 months ago • Mark
Dear DiffBind developers/users,
I have runned my histone ChIP-seq differential analysis using DiffBind some months ago. Back then...Dear DiffBind developers/users,
I have runned my histone ChIP-seq differential analysis using DiffBind some months ago. Back then I saved my output from **dba.analyze(test_contrast, method=DBA_ALL_METHODS)** as an **.RDS file**. At that point I also used DiffBi…
updated 8 months ago • M.Vukic
luck to find a way to mark individual element.
Thank you in advance.
![Figure 11 from the DiffBind document][1]
[1]: /media/images/14f59ed3-ff8e-4ef3-8a78-c39909c7
updated 8 months ago • Junsik
Noticed a discrepancy in config file parameter names.
In all documentation, the DBA$config$edgeR$bTagwise and DBA$config$DESeq2$fitType parameters are structured as written here. However, in the actual DBA object they are stored as DBA$config$edgeR.bTagwise and DBA$config$DESeq2.fitType
This causes problems when trying to run an analysis on a DBA object resulting in the following error:
…
Hi all,
I'm new to ATAC-seq analysis and have recently been trying to use DiffBind to systematically identify differential peaks that I've been seeing by eye when looking at macs2 output in IGV...Hi all,
I'm new to ATAC-seq analysis and have recently been trying to use DiffBind to systematically identify differential peaks that I've been seeing by eye when looking at macs2 output in IGV. …
Enter the body of text here
Code should be placed in three backticks as shown below
```r
# include your problematic code here with any corresponding output
# please also include the results of running the following in an R session
sessionInfo( )
cell <- dba(sampleSheet = "/home/cytolab/extdata/cell lines data.csv")
1 cal27 ko1 H3K36me2 Treated full media 1 bed
Error in if (file.info(pe…
Hi all,
Would you please suggest what wrong in this case? I pretty much run the same code as before. Thank you so much.
result <- dba(sampleSheet=samples)
result <- dba.blacklist(result)
result <- dba.count(result, bParallel=FALSE)
result <- dba.count(result)
result <- dba.normalize(result)
result <- dba.contrast(result, minMembers=2)
result <- …
updated 9 months ago • Chris
Hello,
@rory I have a question regarding the 3 scenarios in which lfcShrink is applied in DiffBind package. The source code looks like this
```
if(shrink) {
if(contrast$contrastType=="byname") {
res$de <- suppressMessages...Hello,
@rory I have a question regarding the 3 scenarios in which lfcShrink is applied in DiffBind package. The source code looks like this
```
…
updated 9 months ago • bhandary.8590
I got the following error while trying to create the initiall dba object:
```r
> tamoxifen <- dba.count(tamoxifen)
Computing summits...
Error: Some read files could not be accessed. See warnings for details.
In addition: There were 12 warnings (use warnings() to see them)
> warnings()
Warning messages:
1: Z:/Shared folder/Shared data/DNA/NGS_AP01_cfrezza_A006200317/A006200317_20…
updated 9 months ago • Theo
I have a TF which is cytoplasmic in control samples & nuclear under treatment and am intentionally trying to demonstrate that TMM is an inappropriate method in this situation. However, I was quite surprised to see that using DBA_EDGER, the results from DBA_NORM_LIB and DBA_NORM_TMM are 100% identical. Fortunately, the expected differences are evident when using DESeq2, but this seems like the…
updated 9 months ago • Stevie Pederson
Hi all,
I got around 100k differential peaks which someone advices me there is something wrong in my ATAC-seq analyses, so I am trying to trace back. Is this sample sheet correct? Thank you so much.![enter image description here][1]
[1]: /media/images/b797f711-4fea-46ec-967c-784dcc55
updated 10 months ago • Chris
We have basically three questions that revolve around the DESeq2 lfcShrink function which is used by DiffBind.
1. We have Cut&Tag samples and want to conduct differential binding analysis. Our main objective is to compare...two samples and see if there are any major peaks that are different. Do you think DiffBind is suitable to conduct such an analysis?
2. In the lfcShrink function th…
updated 10 months ago • bhandary.8590
Hi all, I have 4 samples which are 4 conditions but when I made some plots, I got only 2 conditions. ![enter image description here][1]!
Does this plot mean ws has chromatin more chromatin open (gain) than wf sample? When I rerun the code, I got all 4 conditions but the plot is quite different. There are no gain and loss but only sites. Thank you so much! ![enter image description here][2]
[1…
updated 10 months ago • Chris
I'm using DiffBind for ATAC-seq DA analysis. The MA plot and volcano look nothing like I expect. I do not expect a large difference between...adjusting the offset, the plots look even worse! ![loess_fail][3]
I'm fairly new to both ATAC and DiffBind, so any help and tips would be greatly appreciated!
```
> sessionInfo()
R version 4.2.1 (2022-06-23)
Platform: x86_64-apple
updated 10 months ago • David.Atkinson2
Hi all, I used a different reference genome to align instead of GRCh38 and got this error:
result <- dba.blacklist(result)
Genome detected: Hsapiens.1000genomes.hs37d5
Applying blacklist...
No blacklist found for BSgenome.Hsapiens.1000genomes.hs37d5
No control reads specified, unable to generate greylist.
No intervals removed.
result <- dba.blacklist(result, blacklist = BSgeno…
updated 10 months ago • Chris
Hi! I am trying to run Diffbind for differential enrichment analysis of my ChIP-seq datasets.
To start with, .broadPeak bed files (both ChIP & Input...and .bam files were supplied to Diffbind.
However, it failed to generate the greylist. It is something about my bam files? The bam files are having ensembl chromosome
updated 10 months ago • kyliecode
Would you please help? Thank you so much!
https://bioconductor.org/packages/devel/bioc/vignettes/DiffBind/inst/doc/DiffBind.pdf
Page 22 the manual explains this plot but I am still not sure.
![enter image description here
updated 11 months ago • Chris
to extract such data for specific windows at some point in the analysis. I am asking as I never used DiffBind before and I do not know at which point such control samples are used in the analysis.
Thanks in advance
updated 11 months ago • agata17
Hi!
I use the function dba.report to retrieve differentially bound sites (th = 1)
I found the fold-changes tend to be very small and do not know how to compute them.
For example, at one site
the mean for control is 1.6973 while
the mean for treatment is 4.231,
and the Fold is -0.001057009, p-value is 0.0051515283, FDR = 0.99.
How the fold-changes were computed?
my code is below.
Thank you!
…
Hello,
I am trying to convert fold values obtained from Diffbind table back to expression values (basically unlog it). My question is whether the log used to calculate the log fold...change is log or log2? I looked at the DiffBind manual and it says "The Fold column shows the log fold changes(LFCs) between the two groups, as calculated by the DESeq2
updated 11 months ago • bhandary.8590
Hi everyone, I am trying to use DiffBind to process an unusually large dataset (101 ATAC-seq samples). Is there a way to construct the dba object using `dba()` in
updated 11 months ago • Shashank
Hi I'm rerunning some analysis for ATAC since I've been having trouble validating some results. I have two files I created in March of 2022 using Diffbind I installed through conda, an analyzed DBA object and a report-based DBA object where I set fold=1 and th=0.01
When I...I've been having trouble validating some results. I have two files I created in March of 2022 using Diffbind I installed th…
updated 11 months ago • jandrade
Dear Rory,
Thank you for your efforts in DiffBind.
A basic overview of my dataset. There are two conditions, each having two bio-replicates. I'd like to run differential...be so grateful if you have any ideas. Many thanks in advance!
My code is as follows.
```r
library(DiffBind)
samples <- read.csv("in.info.csv")
tamoxifen <- dba(sampleSheet=samples)
library(rtracklayer)
mype…
Hi all, I am trying to find which genes are different in chromatin accessibility so I used nf-core/ATAC pipeline then feed the bam files and broadpeak files output into Diffbind. I got around 100k peaks, 21k genes after annotated peaks by ChIPpeakanno, and after sorting the top genes by Fold. I...so I used nf-core/ATAC pipeline then feed the bam files and broadpeak files output into Diffbind. I g…
updated 11 months ago • Chris
Hello, I hope you are doing well and are safe. I wanted to reach out since I have been having issues with loading my broadPeaks and/or the .xls outputs from MACS2 into DiffBind. Would appreciate any help you can provide, thank you! My thinking is there might be something wrong with the way I am...since I have been having issues with loading my broadPeaks and/or the .xls outputs from MACS2 into Di…
updated 11 months ago • Mark
Hi ,
I am currently working on 12 ATAC-Seq samples using Diffbind. Following the tutorial, the first step involved generating a dba object and setting the minoverlap parameter...Hi ,
I am currently working on 12 ATAC-Seq samples using Diffbind. Following the tutorial, the first step involved generating a dba object and setting the minoverlap parameter to...A, but when I go back to check the input…
Hi, I've used DiffBind to analyze differential binding analysis for cut&tag data.
The result is showed in MAplot:
![enter image description...Hi, I've used DiffBind to analyze differential binding analysis for cut&tag data.
The result is showed in MAplot:
![enter image description here][1]
I noticed some peaks with 0 log fold change are differentially biding sites, while it…
updated 12 months ago • Ti
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…
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
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.…
updated 12 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
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.
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…
updated 13 months ago • Chris
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