Questions tagged [deseq2]

Deseq2 is an R package for analyzing RNA sequencing data

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deseq2 single factor design output interpretation

This is regarding the single factor design For example if I have Age or other continuous numerical variable how to provide that into the design formula. For this post do i need to 'You could ...
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deseq2 model design : Different gene output

Here I have two designs: Design 1 ...
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deseq2 makeContrasts function

This article talked about various deseq2 designs etc. One of the designs I would like to use is explained as this: Control versus treatment average ...
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deseq2 full and reduced model interpretation

I would like to know how to interpret the output of the formula although i been thorough loads of literature but I'm not confident yet. So this is my full model I'm running ...
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Different results of DESeq2 in different ways of comparison

I have different treatments (infected and control ones). When using DESeq2, I have different results in two ways of comparison. first way: factor: treatment, factor level1: Moribund Infected, factor ...
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What is the difference between Normalized Expression in EdgeR vs DESeq2?

I am trying to access the normalized expression in both edgeR and DESeq2, yet the results are different. Does anyone know why? How to get normalized expression using edgeR: ...
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baseMean threshold

I have an RNA-seq dataset and I am using DEseq2 to find differentially expressed genes between the two groups. I used pre-filtering to remove any genes that have ...
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Error in DESeq2 analysis with dimnames

I'm getting the following error in a DESeq2 analysis and I can't seem to figure out the issue: ...
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LRT test in deseq2 on multiple condition

I have some doubts when i'm running LRT This is my condition data-frame small subset ...
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How to input data and metadata from NCBI for RNA-Seq analysis in R

I am quite new to R programming so I hope my question is a clear one and not too confusing. So, I have downloaded some .txt and .csv files containing the raw count data (I am supposed to find out ...
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How does DESeq2 "collapseReplicates()" function work on read counts data?

Comparing read counts from an RNA-seq experiment for two select genes before and after using DESeq2's collapseReplicates() and ...
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Batch Effects that are Close to Collinear with Main Effect

Two years ago, I started working on a project uses both RNAseq and ATACseq. It's supposed to be a simple Healthy Control (HC) vs disease study. The sample collection was done in 2 phases, and it was ...
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How to remedy a DESeq2 collapsing technical replicates error?

Goal: To ensure "the sum of the counts for [my samples] is the same as the counts in the [samples] columns in ddsColl" after collapsing replicates using ...
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Use of "baseMean" in DeSEQ2

The basemean is described as the "mean of normalized counts of all samples". My question is, how would we interpret differences in basemean between genes, pertaining to reliability and such? ...
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Visualizing top expressed genes

I have a spreadsheet of CPM values I want to visualize the top 20 genes expressed across my samples This is how my data look like ...
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Best R program to use for RNA seq analysis

We have an RNA seq taken from a the same tissue in two different developmental stages. We have one gene, with the number of reads from stage A and stage B, and we want to see which genes are ...
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The correct design for time series experiment

I have a treatment and control in two time points like this ...
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How to simulate replicates for DGE analysis?

I am prototyping with data visualization of DGE results, and I would like to work on the analysis pipeline before the real data is available. Currently, I only have 3 samples for wild type and 1 ...
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Help with performing PCA analysis on data from 3 different datasets of normalized counts data for RNA-seq experiment

So, I have limited knowledge of R but I need to do a PCA analysis of 3 different datasets of gene expression as a result of combined growth or mono-culture growth. The 3 different datasets I performed ...
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Understanding the different designs in DESeq2

I am using DEseq2 and trying to understand the results obtained using different models. I have a data design with 2 genotypes and 2 time points. ...
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Different contrasts in DESeq2

I have a treatment and control in two time points like this ...
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DEseq2 contrast in two time points

I need help with some clarifications please I have control versus treatment in two time points like ...
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Differential miRNA expression using RPM

I have microRNA (miRNA) expression data in RPM. I would like to do differential gene expression between two groups. How can I do this? I have considered edgeR and DESeq2 in R, but it looks like they ...
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Adjusted P-value from DESeq2

What does the padj really mean? I believe it's a BH method, where it attempts to quantify the likelihood of a "false positive" but I am not sure. Also, what does it mean if the resulting ...
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DESEQ2 Question about results()

I am currently learning to perform Differential Analysis via DESEQ2 R Package, and I believe I've made progress, able to format the data correctly [maybe] for DDS(). When I run the results function to ...
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Beginner with DESeq2 having issue with analysis

Hi there I am brand new to using RStudio and trying to run DESeq2 analysis on my featureCounts output counts table. I ran the following code: ...
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DESeq2. Which is better: using the altHypothesis argument in the results function or manually filtering for your desired P-value and Log2FoldChange?

In DESeq2 you can identify your differentially expressed (DE) genes using the results function. I noticed people in my lab using the results() function with the minimum number of arguments supplied (...
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How does FDRtool work?

I have a question about using FDRtool. In the below code (on RNA seq data whose p values were acquired using Deseq2), the FDRtool was first used and thereafter p.adjust using the benjamini hochberg ...
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Calculating most abundant transcript from RNA-Seq data

vcf2maf uses VEP to annotate variants, and I believe selects the default Ensembl transcript to use for annotation. Sometimes the ...
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DESeq2 multiple treatments, multiple time points, multiple cell lines

Note: this question has also been asked on Bioconductor Support I know there are a lot of questions asking similar things here on this forum and I checked the vingette and did a lot of other research ...
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Un-normalize DESeq2 counts

I obtained a matrix of RNA-seq count data that has been normalized by DESeq2's median of ratio method. I know that DESeq2 wants to take in un-normalized counts, but I do not have access to those data. ...
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Bulk RNA-Seq Read Length Normalization across different samples

I have 20 samples out of which 14 are 100 bp in length and 6 are 150 bp. Is there a way to normalize the read length for cross-sample differential expression comparison? What would be the best way to ...
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Differential expression analysis for a subset of transcripts

I want to check the differential expression of a specific class of transcripts (say, long non-coding RNAs) using DESeq2. Now, I know that the normalization step takes into account the total number of ...
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Adding matrix as a confounder in EnrichmentBrowser

Brain data, particualrly human brain data have very heterogeneus cell types, so it's important to normalise by cell type/add proportoins of different cell types to the formula when performing ...
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the model matrix is not full rank : This is a classic question which a biologist face without clear understanding of the model design

Saw this answer biostar. Tried to make my metadata as such but still the "Error in checkFullRank(modelMatrix) : " This is my coldata ...
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What is a sensible number of gene/observations to explain PCA variance?

I am working with a set of RNASeq dataset. I have about 4000 observations (genes) on 20 samples and plotting a PCA I found the clustering doesn't vary much when I use different number of genes, but ...
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Question of Padj value (very high, infinite) got from dds

After I got dds from my count matrix by DESeq(), I use results() method to get Padj and log2 fold changes for volcano plotting, then I found some gene's Padj values are infinite and the P-value is ...
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Why use "robust" FPKMs?

Both DESeq2 and edgeR have an FPKM/RPKM function that by default uses normalized library sizes ("robust" option in DESeq2). FPKMs have their own issues, but I thought the main benefit was to ...
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Is there an established method for comparing expression of groups of genes (gene sets)?

I have a figure in a paper where I show the log2(fold-change) (obtained with DESeq2) between two groups (based on genotype) for genes within a specific hallmark gene set, and a reviewer is asking ...
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Details of DESeq2 modeling a batch effect

When correcting my data for a batch effect using removeBatchEffect, some of the gene expression values become negative. When searching for differentially expressed genes, I do not use the data above, ...
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Setting Contrast for DESeq2 results

This question was also asked on BioStars I am trying to recreate this heatmap. How would I compare the four variables: Ly49+/- and MOG/MOGSP to get a single heatmap? Thank you for helping me through ...
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the variation between treatments is less than the variation between replicates in RNA-seq data

I have a set of RNA-seq samples from targeting different proteins in a complex with siRNAs. However, the ...
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Within and between sample count normalization

recently I came across a situation where RNASeq sample quality was expresses trough DESeq sizeFactor. So authors reported quality of their samples with respect to some publicly available datasets as ...
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Differential Gene Expression with Replicates for some of the samples

[this question has also been posted on Biostars; some additional clarification from there has been copied into this question] I've been asked to analyse a set of samples in which their control sample ...
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a proper Design Matrix for several drug treatments with both control negative and control positive

I have a dataset of RNA-seq samples for testing different drugs on the presence of another drug. One of my samples is the normal cells with no drugs (control negative) and another is the cells with ...
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RNAseq biological replicates not clustering in PCA plots

I have RNAseq data from 4 samples with 3 biological replicates per sample. I am currently trying to do the differential expression analysis with DESeq2 but the biological replicates will not cluster ...
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How to design DESeq2 LRT model with individuals nested in 2 levels?

We have a complicated experimental design that we would like to perform LRT analysis for. Our main goal is to discover significant genes for the "Injection:Social" interaction term across ...
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2 answers
994 views

DESeqDataSetFromTximport all(lengths > 0) is not TRUE

I am suddenly running into an error when running the DESeqDataSetFromTximport ...
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Is it conflicting: significant difference according to Wald test but not in PERMANOVA and ANOSIM to compare species abundances of metagenomic samples?

If there is no significant difference between two groups of metagenomic samples according to "Multivariate differential abundance tests" for example PERMANOVA and ANOSIM, does it makes sense to use "...
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Ranking metric in GSEA

I am using fgsea in r to calculate and plot a bunch of GSEA graphs. My question is whether it is acceptable to use the Wald statistic from DeSeq2 to rank the gene list? I have seen in the GSEA ...
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