Questions tagged [differential-expression]

Use it when comparing a measure of the expression between two (or more) samples, where the units are relative to the comparison.

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11 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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1answer
61 views

Differential gene expression bias due to effect of an individual sample

I am analysing a human single cell RNA seq experiment, where we have 4 groups, four samples each. Data has been analysed using Seurat, with the canonical workflow. I have tried DE using various ...
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1answer
41 views

Seurat define manually a cluster and find markers

I am aware of this question Manually define clusters in Seurat and determine marker genes that is similar but I couldn't make tit work for my use case. So I have a single cell experiments and the ...
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15 views

Filtering criteria for selecting DEGs from Monocle3 regression analysis with categorical terms

I'm testing the package Monocle3 (version 0.2.0) with the following code. The main aim of this analysis is to identify differentially expressed genes for every time ...
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1answer
47 views

How I deal with this expression set?

I have a gene expression raw counts like below for 16 patients ...
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1answer
84 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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1answer
35 views

How to perform DE analysis for each sample

I am new to R and biocondunctor. I have the normalized expression values for 20 samples for a disease and for 10 controls. I wanted to get the differential expressed values for each sample with all 10 ...
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43 views

Identification of differential genes across 8 groups [closed]

I need to identify differential genes across 8 groups. I know this can be done using DESeq2 or EdgeR as these are better suited and equipped. However, I was thinking if something like this piece of ...
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37 views

How could I find Differential Genes in Expression Matrix File by R?

I tried to get gset of my dataset by getGEO function, but unfortunately it didn't work( I installed GEOquery package) and i get errors like this: Found 1 file(s) GSE69606_series_matrix.txt.gz trying ...
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1answer
34 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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2answers
158 views

Strange p-value histogram for differential gene expression analysis

I'm trying to perform pretty standard differential expression analysis using RNA-seq data. I've used Kallisto to perform RNA quantification and am using Sleuth to perform the differential expression ...
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1answer
59 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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50 views

Gene ratio as imput in limma

I have a data frame with gene-expression ratio. Is it possible to input this into limma/voom to find signinficannt gene-ratios between groups of samples? my data: ...
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1answer
74 views

Merging transcriptomes coming from different experiments

I'm planning to build a transcriptome by pooling all existing transcriptomes in SRA for a non-model species (which has no reference genome) to study differentially expressed genes and the like. It ...
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105 views

load the phenotype data for ballgown

I am trying to reproduce the work of this paper [1], and I have run StringTie successfully, but after that I have to run Ballgown but could not understand this command: ...
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22 views

How to retrieve the best-scoring Trinity isoform from blastx results

I'm working on a somewhat unusual transcriptome focused on a killer X chromosome. This chromosome has some new genes, with expression levels quite different from their autosomal paralogs. I've found ...
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1answer
55 views

I need some tips and suggestions for further analysis of NGS expression data (log2cpm)

I am a PhD student who inherited some log2cpm data of expression data from bulk kidney tissue from a UUO(unilateral urethral obstruction) experiment that tests a new drug. The sample material consists ...
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36 views

Common Mouse and human DEG analysis

I have DEGs from human and mouse (equivalent of a human disease model), and would like to generate a logCPM correlation plot between the overlapping mouse and human gene (275 genes). Prior to ...
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1answer
333 views

How to normalise scRNASeq data for differential expression analysis

I wish to perform differential expression analysis for cluster-specific gene expression in single-cell data (with a tool such as MAST or SCDE). I have data for 3 biological replicates. I performed ...
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2answers
925 views

What does an FDR value of 1 in RNA-seq mean?

I am looking at the supplemental data from the paper "An allelic series of miR-17 ∼ 92-mutant mice uncovers functional specialization and cooperation among members of a microRNA polycistron" which ...
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1answer
237 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
271 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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1answer
100 views

Change sample names while using cummeRbund

I want to analyze RNA seq data using R package cummeRbund. Using this package I have a problem changing sample names. When I run this command in cummeRbund ...
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3answers
154 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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1answer
87 views

Is removing samples based on clustering for downstream analysis a right choice?

I'm using TCGA Lung cancer data. I'm interested in doing differential analysis between Lung vs Normal. Before DEA, to check the distance between each pair of samples I plotted an MDS plot: In this, I ...
3
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1answer
102 views

Detecting differentially expressed genes with foldchange >= 2 and FDR < 0.05

I'm using edgeR for differential analysis. Using glmTreat function I'm detecting differentially expressed genes between Tumor and Normal. I have set the arguments like below: ...
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2answers
39 views

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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2answers
108 views

Smallest group size for differential expression in limma (bulk RNA-Seq)

I am reading Smyth et al. (ref. 1). I want to run differential expression analysis on a bulk RNA-Seq dataset in which each group is composed by 2 samples. In the paper previously cited it is written ...
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3answers
240 views

Expression of a gene in different groups

I would like to check the expression of a gene in different groups like Disease vs Normal samples. I want to make a plot out of that to check whether it is significant or not. From this paper lncRNA ...
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2answers
87 views

What kind of analysis can be done with differential expression of transcription factors?

I have two different stem cell types and their respective gene expressions from RNAseq. I noticed that there is differential expression in some of the genes coding for histones as well as ...
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1answer
954 views

Performing t.test to obtain p.values for each gene

I have FPKM values for Normal(80 cases), Group1(135 cases), Group2(147 cases), and Group3(102 cases) for 160 genes (Groups were obtained by clustering tumor data). I want to perform t-test (two-...
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1answer
76 views

How to calculate call detection values in microarray datase if .cel file unavailable

I would like to do a differential gene expression analysis on a microarray data. From the literature, I understood, call detection (Present, Absent, Marginal) has to be done to minimize false ...
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0answers
602 views

Which R package to use for differential analysis with TPM values?

I'm using hisat2, stringtie tools for the RNA-Seq analysis. After stringtie using ballgown I get FPKM and TPM values for every gene. I have seen that edgeR, Deseq2 can be used for Counts data. I ...
5
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1answer
192 views

Which tools for differential expression analysis in scRNA-Seq?

I am starting to run analysis for differential expression in scRNA-Seq. Which tools are available for this kind of analysis? Can tools for bulk RNA-Seq like DESeq be used for scRNA-Seq?
5
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1answer
81 views

Can I use DRIMseq to model interaction effect of variables?

I am trying to use DRIMseq for DTU with 2 treatments on two different strains of mice. In my previous analysis with genes only, I had found a strong interaction effect of strain and treatments. Is it ...
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1answer
321 views

Comparison of gene set enrichment statistics

I am performing a gene set enrichment analysis to determine if particular gene sets are coherently up- or down-regulated. I have seen several statistics for computing a p-value of GSEA-style ...
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1answer
61 views

differential expression multiple studies

I was wondering if we can perform differential expression on samples from different library and read lengths. What we intend to do is, n = 6 tumors (recently sequenced using Hiseq 2500 read length ...
2
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1answer
131 views

Why do I obtain very strange results with DESeq2?

I am using DESeq2 to perform a differential expression analysis, but I obtained very strange results for some genes. For some genes, I have very high log2FoldChange with very low p-value as displayed ...
5
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1answer
122 views

How can I specify to DEseq2 to only perform comparisons on which I am interested with?

I am currently performing a large RNA-seq analysis from mice PBMCs. The dataset contains around 6,000 transcriptomic profiles and I would like to use DESeq2 to identify the sets of differentially ...
2
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1answer
33 views

Simple Affy Differential Expression

I think I have a fairly easy-to-solve problem. I have an expression table generated from an Affy set. It has one row of gene names (already symbols) and then 180 sample rows. 181 rows total. So I ...
5
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2answers
73 views

Differential Expression With 2 Treatments

I have a expression data from a small cohort of samples taken at baseline and after 2 independent treatments. I can do differential expression contrasting T1 and T2 or I can contrast T1 vs baseline ...
3
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1answer
131 views

error in heatmap using R

I am learning about generating heatmaps using R. For this example I am using randomly generated data to test the process, but I encountered what appears to be an error or perhaps there is something am ...
4
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1answer
79 views

Standard Cutoff for Moderated T-statistics

I'm looking at some microarray data. For the first time I've calculated a moderated T statistic from limma. Is there any standard practice for where to cut off that value? For log2 fold change I ...
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3answers
537 views

Error in creating a volcano plot in MATLAB

I am a complete newbie to MATLAB and the first task I have is to create a volcano plot. I have been using this link to understand about it and get started. I tried to run it on dummy values - ...
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2answers
282 views

Correct for gene length or read counts in GO enrichment analysis

It is a well reported fact that GO analysis of RNAseq results is affected by a number of biases, including length bias and expression level bias. The bioconductor ...
4
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1answer
69 views

Using signal peptide and the expression levels of signal recognition particle in secretome analysis

I have not found any work which investigates assessment of differences in levels of secreted proteins by taking advantage of differential expression of the genes which mediate the secretory pathway. ...
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2answers
65 views

Can gene co-expression networks be used to help identify differentially expressed genes?

In my RNAseq dataset of differentiated stem-cell lines, some samples have far fewer significantly differentially expressed genes than others. QC shows that this is because there are way fewer reads ...
8
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1answer
374 views

Sleuth: transcripts with beta close to 0 are considered differentially expressed in a likelihood-ratio test

I'm comparing the results that I obtain when doing a DE analysis with the Wald test and the likelihood-ratio test. One the thing that I've noticed is that there are many genes with 'beta' close to ...
6
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3answers
2k views

Removing PCR duplicates in RNA-seq Analysis

After reading some of the forum posts in Biostar and SeqAnswers I find it very confusing whether to filter out the duplicate reads from aligned files or not. As far I understand it's very difficult to ...
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2answers
356 views

differential gene expression complex design no replicates

We have an experimental design as seen below Where we administered drug at 0 min for each mouse genotype and took them down at given below intervals. wt and ko mouse models were administered only ...