Bioconductor Forum

Hi I'm trying to work with DESeq2, i already worked with edgeR and DESeq1 with relatively sucess. They give me the same results as logFC (+ or -) but, when i work...with DESeq2, i have different results. Every gene who are positive for edgeR and DESeq1 are negative for DESeq2 and vice-versa. DESeq2...I'll post my pipeline, thanks for your help! bckCountTable <- read.table("DF.txt", hea…

DESeq2

updated 7.0 years ago • Luiz

used the following code, and got 266 genes with padj < 0.05: table <- read.delim("test.txt") row.names(table) <- table$Feature\_ID count\_table <- table\[, -1\] conds <- c("ctrl", "ctrl", "ctrl", "treatment") cds <- newCountDataSet(count...method="blind", sharingMode="fit-only") results <- nbinomTest(cds, "ctrl", "treatment") In DE…

DESEQ deseq2 deseq replicates biologicalreplicates

updated 8.5 years ago • wd

<div class="preformatted">Hi All I haven't used LIMMA for a while, but I remember that when I did there was a special way to handle duplicate spots. However the layout of the array had to be such that one replicate was printed first for all spots and then the...I haven't used LIMMA for a while, but I remember that when I did there was a special way to handle duplicate spots. However the la…

limma limma

updated 15.8 years ago • Cecilia McGregor

5 conditions (vehicle, tx1, tx2, tx3, tx4). When I label the groups "A", "B", "C", "D", "E" and run DESeq2 on all 5 groups at once, I get a list of 7000+ differentially expressed genes (padj < 0.05). However, when I label the groups...Shouldn't my results theoretically be the same regardless of how I named the groups, since I gave DESeq2 the same data? Am I doing something wrong/missing so…

deseq2 multiple groups

updated 6.2 years ago • blobbyscience

i have normalized the data using deseq2 internal normalization method. But it seems that my original input data is same as the normalized output. i don't know...on adjusted p value <0.1 Code should be placed in three backticks as shown below ```r library(DESeq2) library(tidyverse) count_data <- read.csv('rawcount_data2.csv',row.names = 1) head(count_data) colData <- read.c…

DESeq2 rnaseqGene Normalization

updated 11 days ago • sajadahmad41454

I have a RNASeq data-set which consists of 4 different conditions (Control, Treatment1, Treatment2 & Treatment3) each with duplicate samples. I would like to identify all genes that are significantly differentially expressed across all the 4 groups...consists of 4 different conditions (Control, Treatment1, Treatment2 & Treatment3) each with duplicate samples. I would like to ide…

deseq2

updated 6.3 years ago • gupta.anuj0608

inspired by this post \#http://seqanswers.com/forums/showthread.php?t=31036 \#install.packages("DESeq2") \#\# try http:// if https:// URLs are not supported \#source("https://bioconductor.org/biocLite.R") \#biocLite("DESeq2") \# Set working directory...head (dat) final\_count <- read.csv("rpkm.csv\_0\_filter\_atleat\_200\_count.csv", header=TRUE, row.names=1) library(DESeq2…

rnaseq deseq2

updated 8.3 years ago • prp291

I downloaded the newest version of R as I kept getting prompted to update in RStudio. As I did, for some reason, previous work I had done following the DESeq2 vignette would not run properly giving me many errors like: 'DESeq2' package doesn't exist. I'll show my most recent errors but I can't run DESeq2 at all. Code should be placed in three backticks as shown below ```r library("DESeq2"…

DESeq2

updated 11 months ago • saguiar

with readGAlignments, I usually set isDuplicate = FALSE  in the scanBamParam argument to remove duplicates. I was just wondering exactly how duplicates are determined--is this based on just chromosome and the start and

genomicalignments rsamtools

updated 7.2 years ago • jfiksel

gt;The duplicated spots have random location, which means that the number of spots between each duplicate is not the same for every gene. This is the summary for the distances: > > Min. 1st Qu. Median Mean 3rd Qu. Max. > 4.00...71.00 86.59 135.00 244.00 > >(Distance here means number of spots between the two duplicates) > >The functio…

limma limma

updated 19.4 years ago • Jason Skelton

vgsID , "GENE.SYMBOL") #then I got Error in `.rowNamesDF<-`(x, value = value) : duplicate 'row.names' are not allowed In addition: Warning message: non-unique values when setting 'row.names': ........ ``` I have use the...duplicate() function to check, but the console return FALSE Does anyone have any idea how to solve this? Thanks

DifferentialExpression

updated 14 months ago • jscl1n22

Hello! I'm working on comparing tumor and healthy samples with DESeq2. My analysis only cares about the adjusted Fold Change values that DESeq2 computes. The input data encompasses a huge...the process up given the fact that I merely require the adjusted fold change instead of the whole DESeq2 output? The full script that I run on the data is [here at this permalink](https://github.com/TCP…

DifferentialExpression DESeq2

updated 6 months ago • Luca

the differential binding analysis of some human histone ChIP-seq samples (bowtie2 mapping to hg19, duplicate marking, MACS2 narrow peak calling, blacklisted regions filtering with bedtools intersect and Anshul Kundaje...with the Y vs H comparison: DiffBind: Error in `.rowNamesDF<-`(x, value = value) : invalid 'row.names' length at the point of my script that I was doing a volcano p…

diffbind

updated 5.0 years ago • melnuesch

Hi!! I'm having trouble to do a contrast using DESeq2. I have 12 samples, AA(2 replicates), AB(4 replicates), BA(4 replicates) and BB(2 replicates). I wanted to compare AB (or BA) with...in resultNames(). Here is my code: <pre> countData <- read.csv(file = dataFile, header = TRUE, row.names=1) colData <- DataFrame(row.names=colnames(countData), condition=factor(c(rep("AA…

deseq2

updated 6.6 years ago • zhuozhu132

gene The duplicated spots have random location, which means that the number of spots between each duplicate is not the same for every gene. This is the summary for the distances: Min. 1st Qu. Median Mean 3rd Qu. Max. 4.00 32.00 71.00...86.59 135.00 244.00 (Distance here means number of spots between the two duplicates) The function duplicateCorrelation in limma can be used to…

limma limma

updated 19.4 years ago • Ingunn Berget

div class="preformatted">Hi , I am using Deseq2 for differential gene expression calculations. I have two conditions Condition a (T) and Condition b (P), when I run the...Thanks in advance, code is : > countsTable <- read.delim ("48_96_filtered.txt", header=TRUE, row.names=1) > pdata = data.frame(condition = factor(c( "T", "T", "T", "P", "P", "P", "P"))) > library …

DESeq2 DESeq2

updated 10.2 years ago • Mubarak hussain Syed

Using the following code to look at differential expression in a couple of datasets. Over and over again the # of up and and down regulated genes is approximately equal. I just wonder if I'm missing something, but I can't find anything about this anywhere. It can't be an artifact of normalization. Any ideas? library("DESeq2") countData <- as.matrix(read.csv("gene_count_matrix.csv", r…

deseq2 gene expression

updated 3.6 years ago • sorrymouse

<div class="preformatted">Hi, I'm trying to use DESeq to know the differential expressed genes of my datasets and i'm encountering that DESeq is not recognizing my row.names so i can't create my cds. My .csv input file looks like: transcript_id,C4,CRL_2APR10,CRL_1_15JUL11,CRL_2_15JUL11 comp1000201_c0_seq1,5.00,0.00,0.00,0.00 comp1000297_c0_seq1,7.00,0.00,0.00,0.00 comp100036_c0_seq1,0.00…

DESeq DESeq

updated 11.0 years ago • Alicia R. Pérez-Porro

permu.size=10000, Pe=0.001) I got this error <pre> Error in `.rowNamesDF<-`(x, value = value) : duplicate 'row.names' are not allowed In addition: Warning message: non-unique values when setting 'row.names': 'cg00042263.ENSG00000146556

HNSC ELMER

updated 5.5 years ago • ghafarpour

div class="preformatted">Hi I want to compare DESeq vs DESeq2 and I am getting different number of DEGs which I will expect to be normal. But when I compare the 149 genes iD that I get...with DESeq with the 869 from DESeq2 there are only ~10 genes that are in common which I don\'92t understand (using FDR <0.05 for both). I want to block the Subject...using:\ \ \ DESeq\ \ library(DESeq)…

DESeq DESeq2 DESeq DESeq2

updated 10.0 years ago • Catalina Aguilar Hurtado

I am posting in regards to an error I've experience when running Deseq2's estimateSizeFactors. The specific error is posted below:  <pre> > dds = DESeqDataSetFromMatrix(countData=filteredDGE...every gene contains at least one zero, cannot compute log geometric means</pre> I've run Deseq2 several times in the past but have never experienced this error. I have trie…

deseq2 estimatesizefactors

updated 8.4 years ago • ssabri

an excellent job with limmaGUI! My question is a simple one. The arrays I work with tend to have duplicates on each slide. However, they are not adjacent nor equally spaced and so when setting the spacing of the duplicates...which appears to be compulsory) I just make it up. I presume this makes the correlation between duplicates meaningless but that linear model fits are OK. Comments? Cheer…

limmaGUI limmaGUI

updated 20.6 years ago • Peter Baker CMIS, Indooroopilly

Hi Rory, I had been previously, not marking duplicates or removing them in my ChIP-Seq datasets, and instead setting the dba.count bRemoveDuplicates to TRUE. Firstly

diffbind dba.count bRemoveDuplicates bUseSummarizeOverlaps

updated 5.2 years ago • loretta

I am using DESeq2 for differential expression analysis. But I got big different results between the two versions while using the same...set of data as input and the same set of r code for data processing for the two DESeq2 versions. I would like to ask what is the key point changed between the two versions to make the results so different...I used DESeq2 package version 1.14.1 and pasilla version…

deseq2 pasilla

updated 6.9 years ago • jingjiaok

I am following this to make pseudobulk and perform DESeq2 https://hbctraining.github.io/scRNA-seq/lessons/pseudobulk_DESeq2_scrnaseq.html Here they perform the DE analysis...colnames(counts) %in% rownames(cluster_metadata))])" for my 3-cell analysis because I can't have duplicate rownames for cluster_metadata. The row names of the metadata are sample names. But the samples would be the same...fo…

pseudobulk DESeq2

updated 3 months ago • puffsquash

<div class="preformatted">Dear List, I had posted earlier about my problem but didn't get any response that could help so I am seeking your help again. I have duplicate spots on my nimblegen array that are position randomized so there is no order to their placement. In Limma when we call the duplicateCorrelation() function, it requires that the duplicate spots have some order to them, lik…

GO limma GO limma

updated 15.0 years ago • Vishal Thapar

Hi, I am using DESeq2 for DE analysis in RNA-Seq experiment. I have 28 CLL patients raw counts and pooled male control and pooled female control...i.e. not matching with existing literature survey). e.g. hsa-mir-155 should be over-expressed but DESeq2 analysis shows under-expressed etc. Here are my codes : expDesign = data.frame(row.names = colnames(cts), condition = c("treated...DESeq…

deseq2

updated 5.4 years ago • vivekr

lt;-read.table(file="TestFile.txt",head=TRUE,row.names=1) expt\_design <- data.frame(row.names = colnames(total\_counts),                  &nbsp...OQ\_YQ.Sig\[complete.cases(OQ\_YQ.Sig),\] write.table(OQ\_YQ.Sig,file="Results\_sig.txt",sep="\\t",row.names=rownames(OQ\_YQ.Sig),   &n…

edger deseq deseq2

updated 6.9 years ago • michael.steffen

Hello Michael, I am new to DESeq2.  I have successfully used the results of function results() to do the plotMA. However, when I try  <pre> lfcShrink...found</pre> My codes are: <pre> setwd("C:/cygwin64/home/Coexpression_Nov2017") getwd() library(DESeq2) library(apeglm) library(ashr) expected_matrix <- read.table(file = "17samples_expected_co…

deseq2

updated 6.5 years ago • lychen83

I tried to follow the [DESeq2 tutorial](https://www.bioconductor.org/packages/3.7/bioc/vignettes/DESeq2/inst/doc/DESeq2.html#how-to-get-help-for...deseq2), and ended up with a error. library("pasilla") library("DESeq2") pasCts <- system.file("extdata", "pasilla_gene_counts.tsv...package="pasilla", mustWork=TRUE) cts <- as.matrix(read.csv(pasCts,sep="\t",row.names…

deseq2

updated 6.4 years ago • unamourdeswann

I have missed any essential components in my script for differential gene expression analysis using DESeq2. I first filtered my data by adjusted p value (0.05) and ranked those by fold change. The following is an example of my output...I be filtering something else? Have I missed something? Here is the R script: ##Load DESeq2 R package library(DESeq2) ##Load in raw re…

deseq2

updated 4.8 years ago • umajs

Hi,  csaw allows one to remove PCR duplicates when doing read counting across windows. My question is how is this functionality actually executed. I generally...mark duplicates using Picard Tools or Samblaster and I was of the view that csaw would remove these reads when doing read counting...wondering if csaw calls some other program (or its own method) internally for marking and remo…

csaw dedup

updated 7.6 years ago • asif.zubair

TRUE) > testFile <- tempfile() > write.table(KarinaSusc, testFile, quote = FALSE, sep = "\t", row.names + =TRUE, col.names = TRUE) > eSet <- read.exprSet(testFile) > eSet Expression Set (exprSet) with 7150 genes 7 samples...sample: arbitrary numbering 2.) Second > KarinaSusc<-read.table("Ka_Susc.txt",header=TRUE,row.names=1,…

updated 20.6 years ago • Marcelo Luiz de Laia

software, specifically with the GUI interface and are running into some problems. Our array has duplicate spots in a top to bottom manner. When we try to have Limma calculate a duplicate correlation we get the error message

limma limma

updated 17.9 years ago • Manish Neupane

followed by featurecounts for raw counts matrix. when i am trying to process the raw_counts using deseq2 the results are not satisfactory to me. please suggest me if it is something wrong with my data![input normalized file...1] this is the r-script which i am running ```r library(DESeq2) library(tidyverse) count_data <- read.csv('rawcount_data2.csv',row.names = 1) head(count_data) co…

DESeq2 DifferentialExpression rnaseqGene DifferentialRegulation

updated 12 days ago • sajadahmad41454

from RPKMs normalized by DESeq2.  I guess this is because DESeq2 reports estimated logfc.  However, sometimes sign (- or + sign) is changed.  Could...res <- results(model, contrast=c("group”,"D16”,"D8")) tmp\_df <- data.frame(res@listData) row.names(tmp\_df) <- res@rownames write.csv(tmp\_df, "./DESeq2\_results/Wald\_D.csv") plotMA(res, alpha=0.…

deseq2 data handling comparing more than two groups normalization logfc

updated 8.5 years ago • bioboy

div class="preformatted">Dear List, I was wondering if there is a tool which can help me remove the duplicate probes based on the location of the probes?. To be precise, I wanted to select the probe which is closest to the 3'UTR

probe probe

updated 11.3 years ago • hsharm03@students.poly.edu

I have repeated sampling of the same individuals I consider it would be appropriate to use Limma and Duplicate correlation (in comparisons involving paired samples) - correct? Q2) We have discussed using Deseq2 as well, but if...I understand, the duplicate correlation function is not available there and therefore it is better to use the code above and limma in the entire...and how and where ca…

RNASeqR

updated 3.0 years ago • sofie.haglund

calculation: featurecount assigned reads/total reads *1000000). Here is an example gene output from Deseq2: ``` Row.names baseMean log2FoldChange lfcSE pvalue padj input_rep1 input_rep2 IP_rep1 IP_rep2 WBGene00006575 70.2292642324974...input2, IP1, IP2, so this small RNA is not enriched in IP. I am not sure if the RPM or log2FC from Deseq2 give the correct result. Here is code I used for Deseq…

RNASeqData DifferentialExpression SmallRNAData DESeq2

updated 7 months ago • Shihui

Hi I'm trying to use DESeq2 to find differentially expressed genes between sample with replicates and sample without replicates. I have 2 condition...control and tumour. I am new to Deseq2 package, do you think my command lines are OK for my experiment?   Thanks in advance   code for find DE in tumour...nbsp; <pre> > countsTable <- read.delim ("/featur…

deseq2

updated 7.4 years ago • Bob

I have run DeSeq2 in 2 different devices using the same count data and metadata tables, however when running the summary and results...both devices and also after round the values I get the same count table values in both, ```library("DESeq2") # sampleTable # csvfile <- "metadata.csv" sampleTable <- read.csv(csvfile, row.names=1, sep=",") sampleTable$Stage <- as.factor...da…

#deseq2

updated 15 months ago • Mafer

I have RNA-seq data from nuclear (Nuc) and cytoplasmic (cyto) fractions of both undifferentiated (UD) and differentiated (DF) cells (I have triplicates of all conditions). Essentially I have UDNuc1, UDNuc2, UDNuc3, UDcyto1, UDcyto2, UDcyto3, DFNuc1, DFNuc2, DFNuc3, DFcyto1, DFcyto2, DFcyto3. I have run DESeq2 on UD nuc vs UD cyto samples and DF nuc vs DF cyto samples separately but I am t…

deseq2

updated 4.1 years ago • AmyNeely2023

https://support.bioconductor.org/p/79209/ and https://support.bioconductor.org/p/98476/). I use DESeq2 in order to get normalized counts from raw TCGA-HTseq Counts. My method is simple, put the data through DESeq2 - using design...all of my analysis (clustering, survival, gene expression heat maps) on the normalised counts from Deseq2. However I am running into problems with huge numbe…

deseq2 limma normalization tcga rnaseq

updated 4.6 years ago • Alex Greenshields-Watson

bioconductor@stat.math.ethz.ch> Sent: Friday, January 07, 2005 1:01 PM Subject: random location of duplicate spots and use of limma > > >> >>Hello >> >>There are approximately 6000 different genes on the arrays...The duplicated spots have random location, which means that the number of >>spots between each dup…

limma limma

updated 19.4 years ago • Ingunn Berget

I could order the genes and get them next to each other. My question is, how would I incorporate the duplicate correlation aspect when defining the design in limma? Thank you for your reply and help. Yolande [[alternative HTML

Yeast Yeast

updated 15.8 years ago • Yolande Tra

nbsp; We perform our DE analysis with limma+voom packages with clean reads without  PCR duplicates removal. for testing how PCR duplicates would affect DE gene and FPKM value, we also map reads with PCR duplicates...DE analysis with the same pipeline. It turns out that most of the DE genes are overlap in both PCR duplicates removal group and without PCR duplicates removal&…

limma differential gene expression bioconductor

updated 8.0 years ago • panying0713

analysis performed by Anota2Seq). I am, however, struggling to understand how to accomplish this in DESeq2. My design is as follows: cts <- read.delim("Interaction.txt", row.names=1) coldata <- read.delim("Sample_Sheet.txt", row.names

deseq2

updated 4.8 years ago • timjoncooper

I realize some areas may map to multiple locations in the genome, but I would like to eliminate duplicates if possible. For example, I have some DMPs that have multiple "hg19_genes_introns" entries. This is a single position...so I would like it to only have one entry. If the number of duplicates were the same for each position, this wouldn't be as much of an issue, because the proportions would …

annotatr AnnotationHub

updated 3 months ago • hayleyw

since I clearly don't know the required R code to do it, I'm here asking for help. I've generated a DESeq2 csv file containing my normalized counts organized by row (Ensembl ID's) as a R df called DSNorm. See my code below. One...be helpful? Thanks all! Code should be placed in three backticks as shown below ```r #load DESeq2 library("DESeq2") #read in the above data cts <- rea…

DESeq2 biomaRt R

updated 15 months ago • Dr.

div class="preformatted">Good morning, I am trying to run DESeq2 with this design formula design <- (~batch+sex+tissue) This is what I do from a count matrix: >library(DESeq2) >countTable...lt;- read.table('matrix.txt', header=TRUE, row.names=1) >ConDesign<- data.frame(row.names = colnames(countTable), batch = factor(c("1", "2", "3"…

DESeq2 DESeq2

updated 10.5 years ago • Ugo Borello

Hello, I am relatively new to using DESeq2 so thank you in advanced for your help! I have multiple samples (A, B, C, D, E) with three replicates of each sample. I would like...Here is the code I'm currently using. ``` ountData <- as.matrix((read.csv("counts.csv", row.names="gene_id"))) colData=data.frame(row.names = colnames(countData),sampleCondition=rep(c("E","E", "E", "F","F"…

deseq2

updated 4.8 years ago • NG

I am doing differential expression using DESeq2. I have 2 conditions (Veh_0h, treatment_56h) and 5 samples each. After generating the gene count matrix from StringTie...I am doing differential expression using DESeq2. I have 2 conditions (Veh_0h, treatment_56h) and 5 samples each. After generating the gene count matrix from StringTie output files and loading into R as `countData`, I used the fo…

deseq2

updated 3.9 years ago • nattzy94

div class="preformatted">Dear all, On my microarray I have some genes for which I have duplicate probes, but not for all. Before I do the stats I would like to average the duplicate probes (e.g. GB10001-RA and GB10006...ideas what would be the most elegant way to average out expression values based on the occurrence of duplicate entries in RG$genes$GeneName? Limma can only average duplicates i…

Microarray probe limma Microarray probe limma

updated 14.2 years ago • Tom Wenseleers

I am using DESeq2 to analysis rna-seq data with 8 biological replicates, which are paired samples. These samples are of primary cells...600px; width:630px"/> 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),subjec…

deseq2 pca

updated 8.3 years ago • g.atla

Hi everyone, I am using ``DESeq2`` (v1.26.0) to find DEGs between ``group1`` and ``group2``, and also between`` group2`` and ``group3``, respectly. And there are ten replicate...will be helpful for me. Here is my code: ``` colData <- read.table('group.txt', header=TRUE, row.names=1) readscount <- read.table('readscount.txt', header=TRUE, row.names=1) condition <- fac…

deseq2

updated 4.1 years ago • Na Chen

that information on following website.[http://bioconductor.org/packages/devel/bioc/vignettes/DESeq2/inst/doc/DESeq2.html#why-un-normalized-counts][1] But, I want to try normalizing raw count into RLE(Relative Log Expression...for your help. ```r # setup condition <- factor(c(c,c,c,t,t,t)) coldata <- data.frame(row.names = colnames(Counts),condition) dds <- DESeqDataSe…

normalization deseq2 StatisticalMethod

updated 14 months ago • SH

Hello, I am new here. I have certain numbers of duplicates for each Probe ID (approx. 45000) in one column and its samples data values in others 'n' columns(approx.. 230 columns

aggregateBioVar DEGseq MicroarrayData

updated 2.5 years ago • biomyscience

Hello all, I tried searching for this specific problem but could not find the answer.  I will happily accept a link to a previous question if such exists. To keep this simple, I can go into more details in response, I am using DESeq2 to analyze the effect of heat stress on expression in two closely-related fish species.  I want to determine if the species vary in their respons…

deseq2 sva interaction term grouping variable deseq

updated 6.3 years ago • kevin.kingsland

<div class="preformatted">Hello, There seems to be a problem with read.ilmn() with the new version of limma. When trying to read the control file, there is an error that row names are not unique. x <- read.ilmn(files="probe-profile.txt", ctrlfiles="JAlverdy-JDeF- Dec2-13-ControlProbeData-FinalReport.txt", other.columns="Detection?) Error in `row.names<-.data.frame`(`*tmp*`…

updated 10.3 years ago • Scott Christley

Hello, I have some questions about running a likelihood ratio test in the DESeq2 package in R. My experiment has two factors: sex and population, with 8-12 replicates per condition. I wish to identify...nbsp; countData <- read.table ("Counttable.txt", header=TRUE) stickleDesign = data.frame ( row.names = colnames(countData), sex = c("M", "F", "M", cont... ) pop = c("A", "A…

deseq2 LRT differential gene expression

updated 8.4 years ago • newsomew13