Bioconductor Forum

Dataset of which I did mapping, and filtered out the genes, and then put it as an expression data in DESeq2, these are some problems I would like to address. 1. when I put in the dataset, I am not able to run it for DEG analysis because...system asks me to remove the first column which is of gene names, and the even if I run it, it gives me only the number in front of the analysis and no name.…

DESeq2

updated 20 months ago • Dev

<div class="preformatted"> Hi, I am currently using the last version of the DESeq2 package to analyse some RNA-Seq data. I would like to use the ReportingTools package to export the results, but I am facing a warning. My code is : > library("DESeq2") > sampleFiles = c("htseqcount_Sample_1.txt","htseqcount_Sample_2.txt", "htseqcount_Sample_3.txt","htseqcount_Sample_4.txt") &…

GO BSgenome BSgenome ReportingTools DESeq2 GO BSgenome BSgenome ReportingTools DESeq2

updated 12.1 years ago • amandine.fournier@chu-lyon.fr

in cell lines we identified genes and exons differentially regulated upon knockdown of a splicing factor using DESeq2 and MAJIQ. Now we want to transfer these findings to TCGA patients with hepatocellular carcinoma (TCGA...problem is choosing the correct normalization strategy for the gene expression estimation. In a first attempt for the 1st task TPM values for all genes in each of the 371 LIHC…

RNA-seq Normalization edgeR recount3

updated 4.6 years ago • mario.keller.1988

I am attempting to run DESeq2, edgeR, and limma-voom to find DEGs between a condition sensitivity with factors 'lineage'. My samples are not really technical

limma DESeq2 RNASeq edgeR

updated 15 months ago • Ben

Hi, I am using Deseq2 to do comparisons between regions of tissue. I have set one region as the the "control" region - BLE. I am trying to pull out...Hi, I am using Deseq2 to do comparisons between regions of tissue. I have set one region as the the "control" region - BLE. I am trying to pull out the pairwise comparisons for the region named BJK which is third in the factor list. I c…

DESeq2

updated 2.1 years ago • claire

the genome annotation does not include isoforms so I don't have a transcript-to-gene map 6. Run DESeq2 pipeline using a single factor (behavior) with four levels. One species has three levels and the other has one level. Here...wondering if there are major problems with the approach that I'm using. I rarely see studies using DESeq2 with more than one species, and I would appreciate any though…

DESeq2

updated 7.4 years ago • fl

the following in an R session sessionInfo( ) ``` Hello, I have a tentative grasp on DESEQ2 I am trying to generate a heatmap (and other DEG visualization tools) using my output from DESEQ2. I followed the tutorial...http://bioconductor.org/packages/devel/bioc/vignettes/DESeq2/inst/doc/DESeq2.html#why-un-normalized-counts) starting with salmon files and then using tximeta to create a su…

DESeq2 DESEQ2 tximeta SummarizedExperiment heatmaps

updated 3.3 years ago • uhlkatie

Hello! I am new to programming and I am trying to analyze some gene expression data using Deseq2. Some of my samples are coming from young animals while some are coming from older animals. For each group I have Disease...see the effect of L-DOPA treatment between different groups but I also want to correct for the age factor if there is any. What would be the best way to do that? ``` SA…

deseq2

updated 6.1 years ago • luca.pagliaroli

First of all I would like to say that I am new to this kind of analysis. I have been using the DeSeq2 vignette extensively and this...have been following the “Group specific condition effects, individuals nested within groups” in the DeSeq2 vignette but I am a little confused with its application.  My data can be substituted in the example vignette (DeSeq2...due to the variance being a …

deseq2 deseq batcheffect

updated 7.4 years ago • bdy8

Hello, I am new to DESeq2. The vignette ([LINK](http://bioconductor.org/packages/devel/bioc/vignettes/DESeq2/inst/doc/DESeq2.html)) focuses on a...that in edgeR we could define such a list via: __my.contrasts <- makeContrasts(). __ In DESeq2, according to the vignette and these topics ([Link 1](https://support.bioconductor.org/p/67600/) and [Link 2](https...ct…

deseq2

updated 7.1 years ago • David ROUX

concentrations (0-120-180-240-300) of 2 molecules (M and S).  I am interested in the 3 factors : Molecule (M or S), Concentration (0-120-180-240-300), Genotype (WT, mut) and I have one blocking factor: Lane (each biological...type <pre> <em>design <- model.matrix(~0+Group)</em></pre> (Group spliting the different factors) would be equivalent to an …

edger deseq2 rnaseq

updated 11.2 years ago • CT

am working through the [PCAtools readme][1]. Two data sets are used to demonstrate the package, the first pca created takes a deseq2 object ([DESeq2 data][2]), and the second does not ([GEO data][3]). The eigencorplot() function is introduced...on the GEO data, and I am trying to figure out how to use it on the DESeq2 data. Can anyone explain why I can't get it to work, and offer a solution? …

DESeq2 eigencorplot PCAtools

updated 19 months ago • BioinfGuru

The first principal component of variance stabilised transformed RNA-seq data for 350 whole blood samples shows some correlation...The first principal component of variance stabilised transformed RNA-seq data for 350 whole blood samples shows some correlation with total read count for sample, and also with library concentration for sample. The PCA analysis is performed on read abundance summa…

RNA-seq PCA control DESeq2 Quality

updated 3.8 years ago • 9906201a

I am using DESeq2 to analysis rna-seq data with 8 biological replicates, which are paired samples. These samples are of primary cells...Here is my code: <pre> x <- read.table("filt_counts.txt", header=T, row.names=1) subjects=factor(c(rep(1:8, each=2))) treat <- as.factor(rep(c("High","Low"),8)) colData <- data.frame(colnames(x),subjects=subjects, treat=treat, …

deseq2 pca

updated 9.9 years ago • g.atla

ATAC_seq_Goett_peaks 11 Samples, 47306 sites in matrix: ID Tissue Factor Replicate Caller Intervals FRiP 1 Cell1px8old epithelial Cell1 1 counts 47306 0.19 2 Cell1_3rd epithelial Cell1...6 mesenchymal 5 5869 6248 and I would like to plot a 4 way venn based on factor. Is it possible? **EDIT** to clarify t…

diffbind plotVenn

updated 6.2 years ago • theodore.georgomanolis

div class="preformatted">Hi Experts, I am interested in predicting transcription factors for a specific family of genes. According to my readings, it is possible to predict transcription factors for the genes

Transcription Transcription

updated 13.6 years ago • Jing Huang

library('DESeq2') countMatrix = read.table("RPR_count.txt",header=T,sep='\t',check.names=F) dim(countMatrix) > head (countMatrix) HRPR_0D_V8_R1...colData <- read.table("Metadata_RPR.txt", check.names=F) dim(colData) # making first column as rownames in coldata SampleInfo <- colData[,-1] rownames(SampleInfo) &a…

deseq2

updated 6.6 years ago • nabiyogesh

Hello: There seems to be a bug in lfcShrink() when using a single factor with more than two levels (three in this case). DESeq() creates a model matrix with 3 columns, while lfcShrink() quietly changes...columns. When I try to provide contrast coefficients to lfcShrink(), it wants to have it both ways. First, it checks if there are exactly three coefficients, then it checks if there are exactl…

deseq2

updated 6.1 years ago • Nik Tuzov

0.1351472 2 10003:106603561 -0.05005979 -0.3629763 </pre> I want to make the first column (rowname) to be my rowname without a column name instead of 1,2,.... I tried rownames(T2LFC) <- T2LFC\[,1\] without success

rstudio

updated 9.8 years ago • John

**Starting context:** I'm pretty intro-level at bioinformatics. I'm using ATACseq datasets. 4 groups (multiple replicates per group) of the following format: Group 1 = ConditionA:TreatmentX, Group 2 = ConditionA:TreatmentY, Group 3 = ConditionB:TreatmentX, Group 4 = ConditionB:TreatmentY **First,** I want to obtain the set of genomic regions that do NOT change in accessibility between…

deseq2 design formula design matrix edgeR ATAC

updated 6.0 years ago • corinne_hutfilz

I am working with a complex dataset design shown here: ![colData][1]. I have followed the DESeq2 manual, but have not been able to find an appropriate matrix model that tests for the effects of Frag_Condition while...tried: ```r countData <- countData colData <- colData # Convert colData columns to factors colData$Site <- as.factor(colData$Site) colData$Colony_s…

DESeq2

updated 21 months ago • kelsey.beavers

Hi, I am using DESeq2 to analyze differential expressed genes (DEGs) from counts generated by Illumina sequencing. I have a 3-factor experiment...Hi, I am using DESeq2 to analyze differential expressed genes (DEGs) from counts generated by Illumina sequencing. I have a 3-factor experiment: * genotype (with 3 levels): A, B, and C, each with 6 biological replicates (3 for each treatment) * …

deseq2 bioconductor rnaseq multifactorial design interactions

updated 9.4 years ago • c.evang

Hello everyone, I am a beginner with the DESeq2 package, and I am trying to understand the mathematical principles behind this package. In statistical modeling...Hello everyone, I am a beginner with the DESeq2 package, and I am trying to understand the mathematical principles behind this package. In statistical modeling and...Where "Y" is read counts and "Beta_1 to Beta_3" is dummy variable…

Regression DESeq2

updated 18 months ago • David W. H.

to do two way anova on the RNA-seq data and find out the variability that can be explained by each factor: <pre> Time <- factor(rep(1:3,4),levels=3:1)</pre> <pre> Sex <- factor(rep(c("Female","Male"),each=6),levels=c("Male","Female"))</pre> <pre> design &lt...model.matrix(~Sex*Time)</pre> I want to calculate the amount of the variance c…

rnaseq two factor anova limma DEseq2 edgeR

updated 9.4 years ago • nooshin

The [DESeq2 paper, page 15](http://genomebiology.biomedcentral.com/track/pdf/10.1186/s13059-014-0550-8) explains the first of the

deseq2 negative binomial model glm

updated 7.4 years ago • jeremy.teitelbaum

interface affect modeling results, within the context of differential expression analysis with DESeq2. I noticed that when I specified two models that from what I understand should decompose the data variance in equivalent...or maybe I missed something. As a toy example, I'm using the same "pasilla" dataset used in the DESeq2 tutorial. The sample meta data is given below. The `group` variable wa…

DESeq2

updated 19 months ago • Ken C

Hello Everyone, I'm currently analyzing a series of RNA-Seq experiments that are splitted into 4 runs. In order to account for the batch effect of each experiments, I'm trying to correct my gene counts using sva, as recommended inside the Bioconductor RNASeq workflow (http://www.bioconductor.org/help/workflows/rnaseqGene/\#removing-hidden-batch-effects) As I proceed in this phase, I got very co…

sva deseq2 batch effect

updated 9.7 years ago • wariobrega

div class="preformatted">Dear Michael, I am using DESeq2 for a factorial design. I have Celltype and condition as colData. I have used the formula ~ Celltype + Cellypte:condition...arises when I want to make an MA plot of the 2 conditions for one cell type. I call the function as DESeq2:: since I known there are some. Here is my code: > ddsFull class: DESeqDataSet dim: 54668 18 exptDa…

GO graph DESeq2 GO graph DESeq2

updated 12.2 years ago • Jose M Garcia Manteiga

I'm currently having some issues with how `` DESeq2 `` implements count outlier replacement when the number of samples suffices. From what I understand, the default is...I'm currently having some issues with how `` DESeq2 `` implements count outlier replacement when the number of samples suffices. From what I understand, the default is to...in the `` A `` group which is below the cooks and is the…

deseq2 deseq rnaseq outliers count outliers

updated 7.1 years ago • lefeverde

I ask for this function, is because I have worked with with the same date in a multifactor desing in DESeq2, where you provide a contrast argument. This is an example from the DESeq2 vignette: resMFType <- results(ddsMF, contrast...my DEXSeqdatasat and code. I have specified two design formulae, which indicate that the StudyType factor should be treated as a blocking factor: > s…

DEXSeq DESeq2 multiple factor design formulaFullModel

updated 10.9 years ago • madal

Day3 Day4 Breed1 3 3 3 3 Breed2 3 3 3 3 We want to use DESeq2 to extract the differential expressed (DE) genes for each time point between the two breeds and the DE genes for each...lt;- DESeqDataSetFromMatrix(countData= milk, colData= design, design= ~ breed + day) dds$breed<- factor(dds$breed, levels=c("breed1","breed2")) dds$day<- factor(…

DESeq2 DESeq2

updated 11.7 years ago • Guest User

Hi All: I would like to ask a question about interaction term in DESeq2. I have the following design: ```r NF2 <- factor(c("Ctrl", "Ctrl", "Ctrl", "KnockDown", "KnockDown", "KnockDown", "KnockDown", "KnockDown", "KnockDown...PRG <- factor(c("Untreated", "Untreated", "Untreated", "Untreated", "Untreated", "Untreated", "Treated", "Treated", "Treated")) prg_metadata_G2G3G5

interaction DESeq2

updated 3.4 years ago • Tait

Hello, I've some bulk-RNA-seq data and I want to perform differential gene expression analysis. This data came from a time course experiment where along a set of time points, wild-type and strain individuals were sampled. I'm familiar with pairwise comparisons in differential gene expression analysis. However, since this data came from a longitudinal study, I think it would be interesting…

deseq2 RNA

updated 5.7 years ago • agsousa

Hi, I've got a question regarding the Cooks distance calculation in DESeq2.  My dataset consists of 4 groups with 5-6 biological replicates per group. For some exploratory QC plotting I performed...Read summarized experiment se <- readRDS(se.path) # Prepare colData group = factor(se$group) batch = factor(se$exp) col.data <- data.frame(row.…

deseq2 cooks distance dds

updated 7.3 years ago • ipso

interested me I noticed a problem. Genes, I assumed to be housekeeping, had decreasing TPM from first to last time point. Some of them DESeq2 detected as differentially expressed. On RPF data these trend was not clear. Also...had significant adjusted p-value. On the other hand, genes with p-value > 0.9 had decrease from first to last sample around 4 fold. Then I calculated rat…

DESeq2 timecourse time

updated 23 months ago • Anastasiia

build networks for gene expression analysis. I hardly get any support from anyone. So I set up my DESeq2 dataset like this: dds <- DESeqDataSetFromMatrix(countData = readcounts_bonobo,colData = coldata_Khrameeva, design...my coldata looks like: ![This is what my coldata looks like:][1] Next I'm going to specify the factors for the comparison. I want to compare this 7 brain area…

DESeq2

updated 2.4 years ago • melchea02

to perform a DE analysis for RNA-seq experiment. I am new to the statistical model implemented in DESeq2 and asking for a help **to write a design formula and extract results using results() function**. **The experiment is the following...timepoints**: KO_1h vs KO_2h or WT_1h vs WT_2h The way I did it was through combining the factors called **genotype** and **time** into a single factor call…

deseq2

updated 6.1 years ago • Kirsan1452

I wonder the best way to analyze such data. I combined Age and condition (genotype) and performed DESeq2 as follows:  > coldata$Age\_condition <- factor(paste0(coldata$Age, "-", coldata$condition)) > dds <-DESeqDataSetFromMatrix

deseq multiple factor design

updated 8.4 years ago • ucheuna

hello all, I want to use p-value of DESeq2 for meta-analysis,I want to know p-value of DESeq2 are normalize?if yes which method DESeq2 use for normalize?  I appreciate

deseq2 normalization p-value metaseq

updated 8.5 years ago • ed_isfahani

to coding on Rstudio. I'm doing a RNA seq analysis to test for differential gene expression using deseq2, by using unpaired samples. I just wanted to know if I altered the following script template correctly to indicate...countdata) head(countdata) # Assign condition (affected versus unaffected) condition <- factor(c("affected","affected","affected","unaffected","unaffected","un…

deseq2 rnaseq

updated 6.3 years ago • adeler001

the mean of a gene across samples 2. Dividing each gene sample by the mean 3. Defining the size factor as the median of the genes divided by the mean But what exactly is the size factor and what is it used for? It is my understanding

deseq2

updated 6.2 years ago • lizzid

Hi, I am curious about prefiltering with DESeq2. I understand from this site and reading the DESeq2 vignette that prefiletering is really unnecessary as DESeq2 has...However, I'm seeing better results with filtering (much higher # DEGs with sig adj p-value after DESeq2). I have a one factor design and three samples in two different groups. I do have a sample that's a bit noisier than the others

DESeq2 prefiltering

updated 3.8 years ago • Chris

Hi Bioconductor community, I performed a categorical analysis (control vs treated) using DESeq2, and I want to put together a figure where I show the average counts in the control and treated groups that includes...Hi Bioconductor community, I performed a categorical analysis (control vs treated) using DESeq2, and I want to put together a figure where I show the average counts in the control an…

deseq2 chipseq counts log2fc error

updated 9.6 years ago • robin.uchiyama

Hello Everyone, I am working on a project with the goal of improving the prediction accuracy of melanoma relapse by combining the RNA signature with the clinical features. Firstly, I calculated the asssociation/relationship between the clinical features with the outcome (early recurrence < 3yrs vs no relapse with at least 5 yrs follow up) and found that Tcat, mitosis and last_vitals…

DESeq2

updated 2.7 years ago • liuyang32201220

two groups at two time points. And the samples are the same in both time points. I have run this in DESeq2: sampleFiles <- list.files(path="/Volumes/timemachine/HTseq_DEseq2",pattern="*.txt"); status <- factor(c(rep("Healthy",26), rep...Normal = unpaired data. I want to compare both within and between samples, so how can I do this in DESeq2? > sampleTable sampleName fi…

limma limma

updated 12.2 years ago • Sindre

tissue) case (right tissue) vs case (left tissue) I would like to find the DE genes present in the first comparison + DE genes second comparison but discarding those DE genes that are significant in both comparisons with...direct way performing only one test and extracting the required contrast. So far I tried this with DESeq2: <pre> design~group</pre> <pre> res <-…

contrast DESeq2 differential gene expression

updated 8.6 years ago • aec

Enter the body of text here When running DESeq2 version 1.28.1, many adjusted P-values have "NA" values even though there are no obvious outliers in the sample transcript...include your problematic code here with any corresponding output I did not get any errors running DESeq2, except for a note on replacement of outliers: > dds <-DESeq(dds) estimating size factors estimati…

DESeq2

updated 4.8 years ago • sagharib

Hi All, I've been trying to wrap my brain around DeSeq2's design model. I was given samples coming from : 2 cell lines, grown in 2 medias, with 2 different vectors. Is there a way...Hi All, I've been trying to wrap my brain around DeSeq2's design model. I was given samples coming from : 2 cell lines, grown in 2 medias, with 2 different vectors. Is there a way to...I've tried a couple of differe…

deseq2

updated 6.9 years ago • jbard

chip-seq dataset which I would like to analyse using DiffBind. I've tried Diffbind with one blocking factor and it works fine, but I was wondering how can I do it for three factors (e.g. age, gender, post-mortem delay , when dealing with...female , 24 hours. ) I ask how to do it because it seems I have to put one of my variables as the "Factor" column in the SampleSheet for the program to easily …

diffbind blocking model

updated 5.1 years ago • melnuesch

Hello, before doing the DESeq2 analysis on my bulk RNA-seq data, I used SVA to identify new covariates and I want to visualize my pca with vst command...in DESeq2, so I did the following passage (I also delete the batch effect of my RIN for the visualization): ```r mm_ALpcrRD <- model.matrix...I have a pca wht 100% of the variance in PC1 and 0% in PC2, do you think it is possible? Is th…

DESeq2 sva

updated 21 months ago • michelafrancesconi8

have to divide into 2 batches for sequencing. Following sequencing, I used Tophat2-featureCounts-DEseq2 pipeline to analyze them.  My question is: should I merge the FeatureCounts result into one file as the input for...DEseq2? Is it OK that I run the pipeline for each batch of samples, generate the DE results (Condition vs control) and then compare...the final results?  __For …

deseq2 featurecounts rnaseq

updated 9.4 years ago • EJ

C 3. A to D 4. B to C 5. B to D 6. C to D but right now using my code I can only get the first three, with A always being compared to. How do I do this? My code is below. `# Assign conditions. I have three replicates in each...group except for the fourth group I'm calling CD49a which has two replicates. (condition <- factor(c(rep("CD103", 3), rep("DP", 3), rep("DN",3), rep(…

deseq2

updated 5.9 years ago • michael_sportiello

we hope to find it is mostly explained by `method`) The way I have been handling this with `DESeq2` is using the design `~ lineage + method`. One of my colleagues claimed, however, that this was an inappropriate use of `DESeq2...that I needed biological replicates that were identical from a design perspective in order for `DESeq2` to be an appropriate choice (eg multiple samples generated fro…

deseq2 differential expression batch correction

updated 6.5 years ago • wunderl

I am trying to run the DESeq2 with only 1378 genes. This subset of genes are defined by the GO term "DNA binding proteins", as I am only interested in...http://seqanswers.com/forums/showthread.php?t=33618). Here are my codes, where cds.dbp is now a DESeq2 object limited to the 1378 genes:   cds.dbp <- estimateSizeFactors( cds.dbp ) cds.dbp <- estimateDispersionsGe…

deseq2

updated 10.0 years ago • meisan406

Hello, This may be a trivial question, but I am trying to use DESeq2 to find dispersion estimates with a reduced model (using no covariates). My dataset has 15400 genes with non-zero expression...per time point. I've emailed the people who wrote ImpulseDE2 and they told me this was possible by first running DESeq2 with a reduced model. Are some genes not able to be used for calculating dis…

DESeq2

updated 4.6 years ago • rohitghosh

I am using DESeq2 to test for differential expression of miRNA reads. The 10 control samples are about 5 to 60 million reads. The 5 treatment...samples are about 1 million to 5 million reads. The normalization run by DESeq2 adjusts these by creating size factors to scale the counts. The overabundance of miRNAs that are found to be down-regulated

DESeq2

updated 4.4 years ago • spollenw

I also read in vignette and many post about multiple factor design but I still don't get it.What I want is to compare between the same cell type and also in the same day but different

deseq2

updated 8.3 years ago • naktang1

Hi, I am analyzing a time course experiment and I am following the example in the DESeq2 tutorial as my experimental design is very similar in that I have two strains sampled at several time points. I would...at any time point and whose expression displays different trends in the two strains across time. At first, I tried doing the analysis the way it is suggested in the tutorial: ddsTC…

deseq2 time series time course

updated 5.3 years ago • bsierieb

<div class="preformatted">Hello everyone, I have been playing with the GLM approach for RNA-seq data in DESeq and edgeR but I am fairly new in DE analyses. I am interested in pairwise comparisons in multi-factor multi-level designs. My question concerns my understanding of an application of the glmLRT function #My code is >countsTable...DESeq and edgeR but I am fairly new in DE ana…

updated 13.1 years ago • Dorota Herman

colData$X8.Session>0,2,ifelse(colData$X7.Session>0,1,0)) #dds needs the design to be as a factor. and associates the 0 and 1 levels with learner and non learner colData$condition1 <- factor(colData$condition1, levels...c(0,1), labels = c("non-learner", "learner")) colData$condition2 <- factor(colData$condition2, levels = c(0,1,2), labels = c("non-learner", "6_7_l…

RNASeqData DESeq2

updated 2.1 years ago • RHEA