Bioconductor Forum

Hi, This is related to the DESEQ2 Normalization. First I would describe my experiment design, I have two groups, GroupA and GroupB. Group A has 39 replicates...sure that the samples from both groups should be included in each batch, therefore, I can use the deseq2 design during analysis. I have run DESEQ2 and run differential expression analysis. I have found around 6700 genes...to me. To exp…

DESeq2

updated 3.3 years ago • MAHESH

I am using deseq2 to find differential significant taxa (count data). The data is longitudinal and three groups of mice and I am using...nbsp;   team + condition:week_experiment)</code> since I have three groups, I wonder how DEseq2 is calculating the fold change and p-value !? There is neither any warning nor any error ! should I slice the data 3 different

deseq2 phyloseq time course

updated 9.3 years ago • akp

I'm trying to do differential expression on label imbalanced data; my case:control ratio is 2:1. I know that regressions are at the core of DESeq2 machinery and I know regressions have internal machinery for coping with such imbalances. Specifically, you can down...on label imbalanced data; my case:control ratio is 2:1. I know that regressions are at the core of DESeq2 machinery and I know regres…

deseq2 rnaseq r regression

updated 9.3 years ago • bkellman

Hello, I have RNA-seq data that I would like to visualize via a GOterm heat map between time points. I have successfully been able to run DESeq2 on my transcriptome, but the output file with logpvalue/pvalue is my entire transcriptome, not compartmentalized by time point (0, 6, 12, 24, 48hr). I was able to make a heat map from this via time point but again, I have A LOT of genes so wanted to gro…

deseq2 goterm

updated 7.0 years ago • shaunnak

Hello everyone, I'm performing a DE analysis between 2 groups of 3 replicates. Samples are *in vitro*-derived, expecting low within-group variance and very (very) high between group differences. Two different cell types are compared and are expected to be quite different. The samples are exosomal small RNAs, which have a very skewed read count distribution. Quite more than typical small RN…

deseq2

updated 6.8 years ago • IV

the number of SVs, only used 2. My question is: should I go with 2 or 9 for the subsequent DESeq2 analysis here? Thanks for any inputs! C

sva deseq2

updated 7.2 years ago • capricygcapricyg

Hi All, In the Deseq2 vignette the first step is creating the object  dds <- DESeqDataSet(se, design = ~ batch + condition) based on the RangedSummarizedExperiment

rangedsummarizedexperiment deseqdatasetfrommatrix deseq2 tximport

updated 8.3 years ago • lirongrossmann

What is the best way to extract the expression matrix, once deseq2 has undertaken it's analysis, if I want to analyse the pre-deseq2 against the post-deseq2 expression matrix using PCA

DESeq2

updated 14 months ago • agamemnon

Hello ! PhD student and working with rna-seq data, I used DESeq2 for my analysis but after finishing my work, I believe some elements were misunderstood when using the software. To...3 effect in G3 The model built with every parameter compared to the references. following the DESeq2 vignette I built the contrast test for example : effect of the T2 compare to the T1 for G1 (treatment 2 ef…

deseq2 R logfoldchange rna-seq

updated 6.5 years ago • pierre.marin

I am having trouble installing DESeq2:   biocLite("DESeq2") BioC\_mirror: https://bioconductor.org Using Bioconductor 3.4 (BiocInstaller 1.24.0), R 3.3.2...var/folders/bt/j1th0gjn1mbdgcr1xh1x3bgw0000gn/T//RtmpmPeLAW/downloaded\_packages > library("DESeq2", lib.loc="/Library/Frameworks/R.framework/Versions/3.3/Resources/library") __Loading required package: S4Vectors..…

deseq2 installation

updated 8.5 years ago • nidz91

We are using DESeq2 to analysis some Ribo-seq data pointwise and try to replicate the dispersion estimation procedure. That is, we focus...count in gene. Thus there is a large proportion of small counts. We have problem to replicate the Deseq2 methods in our data. Could you help us with them? First, following Anders and Huber (2010), we had lots of negative raw dispersions...at small relative…

DESeq2 Ribo-Seq

updated 6.2 years ago • li.keren.cn

Hi, I am reading through the DESeq2 manual and saw the different examples in the results help page. I have a couple of questions to make sure I am understanding...Hi, I am reading through the DESeq2 manual and saw the different examples in the results help page. I have a couple of questions to make sure I am understanding these things properly. I am focused more on "Example 3: two conditions, t…

deseq2 design and contrast matrix

updated 10.6 years ago • Aggarwal, Praful

like described in e.g. the Trinity description into one table for all samples. Striuggeling now in DeSeq2 with the function "deseqtable <- DESeqDataSetFromMatrix" we tried now the tximport package reading the manual but...rounding it first ? Would be helpful if anyone can post some helpful command lines :) 2) is DeSeq2 by default excluding results with low counts (see 1.5.3) - how can I …

RSEM deseq2 count matrix

updated 8.9 years ago • kmeusemann

expressed genes between a wild-type (Ctl) and mutant (Mut). The mutant has a large DNA duplication encompassing several hundred genes, so they always come up as ~2x upregulated but this is not biologically interesting...tt_2 <- run_cf_eb(H0 = 2, fit) ## H0 is 'ctl = 2 x mut' ## tt_all <- merge(tt_1, tt_2, by = 'row.names', suffixes = c('.H0_1','.H0_2')) ## Plot logFC from H0('ct…

differential gene expression limma rnaseq design and contrast matrix limma voom

updated 9.9 years ago • stuart

Hi, I like to use DESeq2 for making PCA-plots and heat maps, but on a current dataset we only have count values from cufflinks/cummeRbund (exported...using count() in cummeRbund). I know DESeq2 needs raw counts, but can I use these counts only for plotting/visualization? And can I perform the rlog-transformation

deseq2 cummerbund cufflinks

updated 10.4 years ago • Jon Bråte

**Background** I am analyzing RNA-seq reads to look at transcriptomic response to environmental stress, and I want to ensure that I am using the correct design formula for my experimental design. I have a condition factor with a control level and four other levels in no particular order (CL, A, B, C, D), biological sampling was done in triplicate, and there is a timepoint factor with four orde…

deseq2

updated 6.9 years ago • galen.seilis

Hi All, I am using DESeq2 and I want to use GC content, gene length and size factor offsets using CQN package, as I do various QC visualization...Hi All, I am using DESeq2 and I want to use GC content, gene length and size factor offsets using CQN package, as I do various QC visualization plots and statistics, I have been trying to export the data with varianceStabilizingTransformation. First …

deseq2

updated 9.1 years ago • kodream

single_pat = read.table( "/home/alva/AML_patients/BM_ID_Rel_CR/ID_REl_CR_BM_counts",header=TRUE, row.names=1 ) head(single_pat) single_patDesign = data.frame(row.names = colnames( single_pat ),type=as.factor(c("patient_4","patient_5

deseq2

updated 11.2 years ago • alva.james

Hello! I am doing DESeq2 for the first time. A bit of background: I am downloading the already public data available from ENA browser. I have been...Hello! I am doing DESeq2 for the first time. A bit of background: I am downloading the already public data available from ENA browser. I have been able to successfully do Kallisto on the paired reads. The output of such files in in .tsv format. I…

DESeq2 Kallisto

updated 23 months ago • Aaliya

Hi! I am using DESeq2 on an in-house server which is shared among a handful of users and has not a cueing system. I want to set a limit on the number...of nodes DESeq2 uses, for which I used `MulticoreParams` as follows: ```r parallelParams <- MulticoreParam(workers = 4,progressbar = TRUE...bpforceGC: TRUE bplogdir: NA bpresultdir: NA cluster type: FORK ``` However, whe…

DESeq2 BiocParallel

updated 3.2 years ago • Sebastian

have a counts matrix of K9me2 enrichment at each gene. I would like to use this counts matrix in DESeq2 for differential enrichment analysis, as if treating it as RNA-seq in essence. However one issue I have is that for Cut...This means my data table now contains spike-in normalized values and not raw counts, which I know DESeq2 normally requires for DEseq(). If I round my values so that no erro…

deseq2 Cut&Run

updated 7.1 years ago • MPerch

I'm running a deferential expression pipeline that first calculates DE with DEseq2 and then looks for enticement in the DE genes. This pipeline seems somewhat common in the literature, so I'm sure...I'm running a deferential expression pipeline that first calculates DE with DEseq2 and then looks for enticement in the DE genes. This pipeline seems somewhat common in the literatur…

deseq2 goseq gene ontology differential gene expression

updated 8.4 years ago • david.rinker

Dear community, I'm trying to use DESeq2 to test if there is a relation between the expression of any genes and X (a continuous variable). For my design I have 5...Dear community, I'm trying to use DESeq2 to test if there is a relation between the expression of any genes and X (a continuous variable). For my design I have 5 persons

deseq2

updated 9.1 years ago • sasha

of both padj and fold change ratio to define differentially expressed genes (DEGs) identified by DESeq2. Would it be valid to submit a manuscript where the DEGs are identified only with a padj threshold? Many thanks for the

deseq2

updated 5.4 years ago • marija.buljan.2

Hi,  I am trying to get direct dispersion estimates from DESeq2. As the results() does not contain the direct estimates, is there an way to get hold of the estimates ? I want plug in these

deseq2 dispersion R

updated 9.5 years ago • hiraksarkar.cs

I am trying to understand some results from DESeq2. The experiment is as follows: Before and after treatment for 11 individuals. The most significant gene has the following...only, except from the second individual, who has 1298 counts before treatment). The p-value from DESeq2 is: 1.49e-21 (using the default Wald test). Here I have used the DESeq vignette instructions for doing paired comp…

deseq2

updated 6.7 years ago • staaln

Dear Michael, I am hoping to use DESeq2 to analyze the sequencing results of a Multiply Parallel Reporter Assay that I performed. I was wondering if you might...that DESeq originally calculated dispersion for each condition separately. My question is: __Does DESeq2 still calculate dispersion for each condition separately, such that the low dispersion of my plasmid reps will not

deseq2

updated 9.7 years ago • dsg16

I am trying to run a time course DESeq2 analysis with 16S amplicon data in a phyloseq object. All treatments in the time course have the same starting point...others and must be removed. Please read the vignette section 'Model matrix not full rank': vignette('DESeq2')</pre> [Michael Love has previously suggested a remedy for model matrix rank issues with time series](https://s…

deseq2 phyloseq multiple time points

updated 8.5 years ago • campbell87

Hi all, a theoretical question about DESeq2. I have a dataset of sorted blood immune cell types from healthy and diseased humans. I want to calculate DE genes for...have different amounts of RNA (e.g. neutrophils vs B cells, etc.), the normalisation method that DESeq2 uses internally might give me erroneous results. Do you think I can still use DESeq2 or should I use a different package

deseq2 normalization

updated 6.1 years ago • tkapell

The documentation of the DESeq sizeFactors, as described in the supplement to the DEXSeq paper says: "we multiply them with a suitably chosen constant c ... such that \[product of final sizeFactors\] = 1. This keeps the size factors close to unity and ensures that normalized count values ... remain close to their original values, making their interpretation, e. g. in plots, easier." But I find …

deseq2 sizeFactors

updated 8.1 years ago • katzman

Hey Mike, a couple of questions on DESeq2, but first of all, some code to make my questions reproducible: <pre> library(airway) library(DESeq2) library(magrittr) dds_airway...lt;- DESeq2::DESeqDataSetFromMatrix(assay(airway),                &nbsp

deseq2

updated 9.2 years ago • Federico Marini

from this, I have 3 overlapping timepoints - baseline, d1, and d2 timepoints. My understanding of DeSeq2 is that as long as the biology is maintained (that is to say that these samples were treated the same), then the more information...I feed into it, the more accurate a model it can generate. So I am inclined to feed Deseq2 all of the timepoints, but perform differential expression analysis li…

Deseq

updated 3.8 years ago • Alex

I am trying to teach my students the using of DESeq2, and they are repeating the instructions contained into the vignette  And they have found a very small bug. This...I am trying to teach my students the using of DESeq2, and they are repeating the instructions contained into the vignette  And they have found a very small bug. This bug does not appear when the "old" DESeq is i…

software error

updated 9.2 years ago • arfranco

Hi,  I have a question about how DESeq2 estimate dispersion parameters. In the low level function of DESeq2, when estimating dispersion alpha, I think the

deseq2 dispersoin estimatedispersions

updated 7.7 years ago • xr2120

readcount matrix form TCGA. The size is 577 samples with number of genes 18.522. When I tried to run DESeq2 to calculate log foldchange, it took not that long, around 3-4 hours. After that, I want to use rlog function to get the log...Core™ i7 CPU 975 @ 3.33GHz × 8 with RAM 24 GB. I know that R can not use multiple core to calculate DESeq2. Is there any suggestion how to optimize this process

deseq2

updated 9.9 years ago • bharata1803

This is my first time using EdgeR or DeSeq2. I used HTSeq-Count to get counts for reads aligning to CDS (this is for ribosome profiling) for each gene. I obviously...This is my first time using EdgeR or DeSeq2. I used HTSeq-Count to get counts for reads aligning to CDS (this is for ribosome profiling) for each gene. I obviously have...reads and reads that don't align to the feature (CDS). Should …

edger deseq2

updated 7.0 years ago • cottrellka

Hi, I am using DESeq2, but unsure whether or not I am using the right design. I could not find here anything similar. **subject**: A, B, C, D, E, F **cohort...Hi, I am using DESeq2, but unsure whether or not I am using the right design. I could not find here anything similar. **subject**: A, B, C, D, E, F **cohort**: ctrl...ctrl or treatment) and each subject is sampled at mult…

deseq2 nested de

updated 6.1 years ago • gaio.transposon

Dear folks,     I got stuck about how DESeq2 get inference or calculated test\_statistics for differential tests. There is a gene of my interest. I want to dot testing between 2 samples/conditions (no replicates) to see if it is significant differentially expressed. Here is the normalized count for them:   …

deseq2

updated 10.3 years ago • xfwang

May I clarify a part of the DESeq2 package? Is it true that DESeq2 counts for linear (only) transcripts and excludes any possibility in circular RNA (circRNA...decrease in circRNA has no effect on the number of counts produced in the results/output? Or does DESeq2 not able to distinguish the two, will assume circRNA as linear fragments, and include it into the counts

deseq2 circrna rsubread

updated 7.5 years ago • csijst

Hello, I usisn DESeq2 package in R for analysing Rna seq sngle end data. After running the DESeq pipeline dds <- DESeq(dds) I am getting this

deseq2 Biocodunctor

updated 6.2 years ago • aapapaiwannou

Hi, I am analyzing RNA-seq and want to put cqn offset (GC% normalization) to DESeq2 object. I followed the vignette of DESeq2 but have a question about geometric mean standardization of cqn offset. ```r

cqn DESeq2

updated 3.4 years ago • bioinfo

all together in the count matrix and later contrast to specify the sample vs  samples in DESeq2                        &nbsp...samples) in the count matrix and compare all Knock-…

deseq2

updated 8.8 years ago • deena

Hi all  for prevent p-values set to NA in DESeq2, should I offset my raw reads as same as this ? Thanks <pre> cont1 cont2 cont3 tumor1 tumor2 tumor3 geneA 64 80 36 0 0 0 geneB

deseq2

updated 8.2 years ago • Bob

Hello! I'm using DEseq2 to perform differential gene expression analysis between two conditions. When I normalized each group separately...Hello! I'm using DEseq2 to perform differential gene expression analysis between two conditions. When I normalized each group separately I find that one of the conditions has significantly larger dispersions in most genes. How can I preform a DEseq2 anal…

DESeq2

updated 2.4 years ago • adi.rotem

Hi all,   We are performing a transcriptome analysis with DESeq2. We have three factors: treatment (control and drought), climate (CO2 and ambient) and zone in the leaf (zone1, zone2, zone3...Hi all,   We are performing a transcriptome analysis with DESeq2. We have three factors: treatment (control and drought), climate (CO2 and ambient) and zone in the leaf (zone1, zone2,…

deseq2

updated 7.9 years ago • Jonas B.

Hi, I'm using DESeq2 for DE analysis. I found in the "Beginner's guide to the "DESeq2" that it's possible to perform both default independent

deseq2 INDEPENDENT FILTERING

updated 11.3 years ago • Laura.Fancello

realised it probably belongs here. Sorry about that. I have a table of FPKM values generated by DEseq2, and I'm trying to find out what DEseq2 uses as gene lengths when these are not supplied (I'm trying to assess to what extent...a given row of object, e.g., the union of all basepairs of exons of a given gene." Does that mean DEseq2 directly uses the ranges obtained by using rowRanges(dds…

deseq2

updated 5.8 years ago • hasse.bossenbroek

Hello there, I guess I would like some clarifications on how DESeq2 handles multiple testing in terms of contrasts of interest along with contrasts of other variables. For example...Hello there, I guess I would like some clarifications on how DESeq2 handles multiple testing in terms of contrasts of interest along with contrasts of other variables. For example, please...It looks to me that…

deseq2

updated 5.7 years ago • g.wang2

Hi, We are currently using the DESeq2 R package for analysis of differential expression of small-RNAs over genes and we would very much appreciate if you...also affect and shift the results. This leads me to the question itself: with the intention to use DESeq2 for differential expression analysis, which counting method would you recommend to use while dealing with small

rnaseq deseq2

updated 10.6 years ago • lehouri

Hi,   I am using version 1.16.1 of DESeq2.  According to the documentation, a Cook's distance cutoff of 0.99 is applied to the results function by default...1.16.1 in Bioconductor up to date with the source code on github? (https://github.com/mikelove/DESeq2).  I have not done much digging otherwise.    Thanks,   Will

deseq2

updated 7.5 years ago • wpoehlm

Hi, I'm trying to do pathway analysis for DE porcine genes generated using DESeq2.  (A) I used package "clusterprofiler" to convert gene symbols to enterzids using the following code: <pre> gg = bitr(m1...BgRatio pvalue p.adjust [7] qvalue Count <0 rows> (or 0-length row.names)</pre> I was able to get a list of KEGG pathways usi…

reactomepa clusterprofiler

updated 7.8 years ago • thomasjenner333

Hi there! I'm interested in DEG analysis in RNA-seq and I have a question about statistical analysis methods. I tried to analyze DEGs using three different methods as follows: 1. DEseq2 package 2. FPKM t-test in excel =T.TEST(ctr1:ctr3,trt1:trt3,2,2) 3. log2(FPKM) t-test in excel =T.TEST(ctr1:ctr3,trt1:trt3,2,2) Actually, I'm a bit confused between 2 and 3. I feel like I should do 3 to a…

DESeq2 statistics rna-seq

updated 2.7 years ago • SH

to one from package ‘S4Vectors’: not turning on duplicate class definitions for this class. Note: the specification for S3 class “xtabs” in package ‘IRanges’ seems equivalent...Annotated” in package ‘IRanges’ seems equivalent to one from package ‘S4Vectors’: not turning on duplicate class definitions for this class. Note: the specification for class “DataFrame” in package ‘IRanges’ seems eq…

org.hs.eg.db

updated 10.4 years ago • alok.jaiswal

Hello all, I want to ask a question about Log2FoldChange in DESeq2. In many cases for the similar counts of Case and Control if the count shows down regulation but the DESeq2 give the output...table __xyz1-xyz6__ results are Down Regulated but the DESeq2 give the Up Regulation. where xyz7-xyz10 shows the correct result. Please help me in …

deseq2 R log2fc

updated 10.1 years ago • Sushant Pawar

For the TCGA COAD HTSeq count data matrix from UCSC Xena, what is the best way to input it into DESeq2 to do DE analysis at gene level? The data matrix is 60488x513, which seems to be not at gene level. Right now, DESeq2 vignette

deseq2 TCGA HTSeq-count

updated 5.5 years ago • zhaoy

After the DESEQ2 analysis, the baseMean values from the results range from (0.2 to 87622.00). For the selected results I have used 0.05

deseq2

updated 6.9 years ago • angajalaanusha

Hi all I'm working with DEseq2 on RNAseq data and I observe an excess of positive signals (see attached MA plot). Any idea about how to reduce the noise

deseq2

updated 6.8 years ago • mesaez

Dear all, How can I filter the  counts with low count in Deseq2?  Any suggestion on how to do? <pre> dds<-DESEq(dt) count<-counts(dds,normalize=TRUE) filter<-rowsum(count)> 10</pre

deseq2 lowcount

updated 4.9 years ago • aristotele_m

T3 1/3 (T1 + T2 + T3) = T4 1/4 (T1 + T2 + T3 +T4) = T5 How would I implement this contrasts in DESeq2

deseq2

updated 8.8 years ago • yohannesafew

Hi I would like to exclude lncrna and pseuodogens from DEG results (using DESeq2). At what step do I do this? Thanks

filter deseq2

updated 2.7 years ago • PD