Questions tagged [deseq2]

Deseq2 is an R package for analyzing RNA sequencing data

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18 views

Differential Gene Expression with Replicates for some of the samples

In some specific situations, it is neither affordable nor feasible to have replicates for large RNA-seq analysis. Moreover, the analysis purpose is just hypothesis generation for further experiment ...
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23 views

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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44 views

How can I perform differential expression analysis with DESeq2, starting with a count matrix?

I have a gene expression count matrix with the Gene Symbol IDs as the row headings and the Sample IDs as the column headers. How would I use DESeq-2 for the given data set through R?
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57 views

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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57 views

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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74 views

DESeqDataSetFromTximport all(lengths > 0) is not TRUE

I am suddenly running into an error when running the DESeqDataSetFromTximport ...
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1answer
28 views

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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48 views

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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1answer
42 views

How to incorportate RIN values as covariate in the design matrix?

I have been following the last DESeq2 pipeline to perform an RNAseq analysis with a dataset with low rin samples in the experimental (or treated) and high rin on the control ones. I read a paper in ...
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1answer
52 views

Should biological replicates be the most similar pairs in an RNAseq experiment?

The attached figure (from deseq2) shows the sample to sample distances of a RNAseq experiment with 4 conditions (A,B,C,D) at 2 time points (0h,4h) with 2 biological replicates. I am a little bit ...
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32 views

Identify differentially covered genes only between two samples

I have a question about finding differentially covered regions (coverage represents methylation level which goes from 0 to several thousands). I'm using enrichment based method which can be summarized ...
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vst() from DESeq2 vs voom() from limma

I have used both to transform my leukaemia RNA-Seq data for subsequent hierarchical clustering. The result is quite different. Some subtypes of leukaemia only form a cluster (or at least sit closer) ...
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25 views

DESeq2 compare within a condition

This might be a really stupid question, but I can't figure out how to do this. I've read through the DESeq2 vignette and manual pages but couldn't find an answer. I have a bunch of samples split up ...
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32 views

Is there a computational tool or possibility to identify mRNA isoforms from the count matrix of a bulk RNA sequencing dataset?

I have the counts matrix of an RNA sequencing dataset of fibroblasts and I wish to identify isoforms of a particular gene of interest in it. Can anyone please hint me on a bioinformatics method to ...
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64 views

How do I differentiate outliers from in-group variations in a DESeq2 PCA plot of 9 samples distributed into 3 conditions?

I have a PCA plot from DESeq2's plotPCA(vsd, intgroup=c("conditions")) function. I have 9 samples distributed in to 3 groups of 3 biological replicates each. My ...
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46 views

How are these definitions related to differential expression?

I have two groups of patients; for each patient I have an output file (RNA-seq) contains this information ...
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559 views

STAR quantMode vs RSEM vs Kallisto

I recently discovered this Snakemake pipeline for RNASeq that uses STAR's quantMode to quantify gene expression for DESeq2 differential expression analysis. In the past I've always seen the workflow ...
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28 views

Comparing factor levels in deseq2

I have sample information (sampleinfo) in this format: ...
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87 views

RNASeq: Normalization, stabilization, gene length and rlog

I was thinking about the best method for normalization, which takes gene length into account (in order to compare genes)... Do you think I can do that? : - taking raw counts and dividing each gene by ...
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1answer
85 views

How I deal with this kind of gene expression comparision

I have RNA-seq from two sequencing batches; Lab technician says that he has run the RNA expression quantification two times in bathes 1 and 2 for example tumor 1 in batch 1 and tumor 1 in batch 2 , ...
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182 views

Interpreting this PCA plot for RNA-seq

I have RNA-seq from two sequencing batches; Lab technician says that he has run the RNA expression quantification two times in bathes 1 and 2 for example ...
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91 views

Comparing RNA-Seq and Ribo-Seq

I'm attempting to compare total RNA-seq with Ribo-Seq, to determine if changes in Ribo-Seq are due to changes in transcriptional expression (akin to the analysis performed by Anota2Seq). I am, however,...
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128 views

Correlate DEGs from DESeq2, EdgeR and Limma results

I have a lists of DEGs identified by DESeq2, EdgeR and Limma. I would like to correlate the the gene rankings in the lists to decide on a package to use in downstream analysis. I am havig a few ...
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1answer
50 views

Single sample in group: normal pipeline or Kal's Z test

As stated, which one is better for differential expression analysis? When I say normal pipeline I mean limma-voom, edgeR and DESeq2 pipeline for standard analysis. Kal's z test is mentioned in this ...
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69 views

design formula question

I am not sure I am building the proper design formula for the question I want to answer I have the following samples with three factors; clone, the structure and the condition. ...
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167 views

Why does DESeq2 convert numeric columns to factor during differential expression analysis?

I'm attempting to perform differential expression analysis using DESeq2 on a dataset of ~90 samples and ~20000 transcripts. I have a number of variables I'd like to test against, some of which are ...
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234 views

Is it advisable to remove X and Y chromosome genes in a bulk RNA-seq dataset at the level of the count matrix?

From this link remove X and Y chromosome genes in RNA-seq data using DESeq2 pipeline, I have learned that depending on context, it is perfectly valid to remove X and Y chromosomal genes in an RNA-seq ...
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90 views

How to get log2 fold change of RNA-Seq data for time series experiment?

I know if there is one control and one treatment group it is pretty straight forward to interpret log 2 fold change. But, I have time course experiment. I have infected cells with viruses and I ...
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183 views

Differential expression analysis when nested effects

I have 3 tumour samples from 3 patients from an experiment, I also downloaded 10 normal samples from TCGA. My design is like this ...
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38 views

Finding genes especific to microenvironment

I have RNA-seq .bam files for 3 patients, tumour and its matched derived model, namely organoid, but I don't have any matched normal sample. Differentially expressed genes between a tumour and its ...
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888 views

Installing DESeq2 in Ubuntu

I am trying to install DESeq2 in my Ubuntu with R version 3.5.1. According to the package repository in Bioconductor the version should be 3.5. ...
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167 views

Removing genes with less than a correlation cut-off between two matrices

I have two matrices like this: ...
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59 views

Making a model of gene contribution in cancer diagnosis

I have raw read counts (Edgeseq technology) of 56 patients who have been ranked with Mandard score as responders and non-responders; I have done differential expression by DESeq2 and I obtained a ...
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69 views

Experimental Design for Differential expreression analysis

I have a Normal esophageal Fibroblasts (NOFs) cultured in DMEM media; The same NOF also have been cultured with a tumor sample from a patient named 005 on DMEM media; I have also Cancer Associated ...
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209 views

PCA plot shows big difference but not many differentially expressed genes are found

I got a PCA plot of bulk RNA-seq experiment that looks the following way: It was generated by the following code: ...
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1k views

Install DESeq2 through anaconda

I tried running conda install -c bioconda bioconductor-deseq2 in a conda environment, but when I run ...
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509 views

DESeqDataSetFromTximport invalid rownames length

I am trying to use DESeqDataSetFromTximport function from DESeq2 package to construct dds ...
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360 views

levels of factors in the design have non-unique level names after make.names() is applied

When I try to import the data using DESeq2 package: dds <- DESeqDataSetFromTximport(txi, sampleTable, ~Group + Cre) I am ...
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1answer
648 views

How to get the corrected matrix after SVA batch effect correction

I ran SVA to remove batch effects for my bulk RNAseq experiments, but now I need to somehow correct my data matrix in order to ...
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1answer
276 views

How to input data for DESeq2 from individual HTSeq count?

I am comparing the gene expression of 2 bacteria under 1 condition. I have now the count tables for 3 tech. replicates for each bacteria. ...
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285 views

Compare clusters from different datasets

I have two scRNAseq datasets, and I select a cluster from one of them, and another cluster from the other. I want to find differentially expressed genes between the ...
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1answer
508 views

DESeq2 complicated design - effect of replicated samples

I have RNAseq data from a relatively complicated experimental design with variables = genotype, treatment, time, and batch. I have 2 biological replicates for each genotype/condition, however ...
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3answers
186 views

Selection of differential expressed genes

I'm working with RNA-seq data. I have 40 tumor samples and 5 Normal samples. Differential analysis with Deseq2 based on Fold change > 1.2 and alpha < 0.05 gave ...
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2answers
1k views

gene-level versus transcript-level analysis

Traditionally, RNA-seq data was quantified on gene level. Newer methods quantify on transcript/isoform level. For example, Kallisto only outputs transcript-level abundances. From the DESeq2 vignette: ...
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Can I ask DESeq2 which variable alone explains which gene's behavior the best?

Imagine I have a dataset of highly periodic gene expression, taken at densely spaced timepoints. I know that the expression is periodic, but different genes have different offsets. It is trivial to ...
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454 views

Likelihood Ratio Test in DEseq2

I have a RNA seq data which I am trying to identify DEGs. Dataset contains: Untreated - 2 replicates (time point 1st day) Treated - 4 replicates (2 replicates at 3rd day & 2 replicates at 7th ...
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1k views

Transform and feed data into DESeq2 with DESeqDataSetFromMatrix

There is a normalized expression matrix. I split it into two and want to do DE on the two cells' subsets. I am having trouble ...
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73 views

differentially expressed intron analysis?

I have used Deseq for estimating the differentially expressed introns for the RNA seq data using the UCSC intron bed file. can Deseq be used for differentially expressed intron analysis?
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43 views

Deseq2 output issue

3 Control replicate and 2 test sample ,so Im comparing my Control which is my Progenitor cells to Monocyte which is mature stage this is my code im using ...
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975 views

Highly variable genes analysis by DESeq2

In the following article. There are two ways of getting highly variable genes: ...