Questions tagged [limma]

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Problems in conducting batch effect correction

I am now trying to conduct batch effect correction for my data matrix, whose samples have different gender and come from different sequencing platforms. I have tried ...
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How to use the contrast function in ebayes for making different comparisons

The design matrix i have is this I would like to know how to use they way its done in deseq2 where we can use the contrast function to make particular comparison The code im running to make ...
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2 votes
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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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Generating contrast matrix for limma in loop

I am trying to generate contrast matrix for different conditions using for loop. ...
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50 views

Extracting adjusted expression after adjusting for covariates

For a given number of features (hundreds ~ thousands protein quantifications) and samples (hundreds) I am fitting a multi-linear model where I am aiming to regress out the effect of a few covariates. ...
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1 answer
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Help with Limma-model

I'm starting to use Limma to find differential expressed sites within my data and would like to ask if my approach was correct. The data has a control and a treatment set, each containing data of 48 ...
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Assumptions of batch effect removal

What does removing a batch effect (e.g. with limma::removeBatchEffect) assume about the batch effect? Does it assume simply a constant batch effect for each level? ...
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1 answer
135 views

Limma decideTests function: what kind of multiple hypothesis testing correction does parameter "method" involve?

What kind of multiple hypothesis testing correction does method="global" do in Limma's decideTests function? According ...
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3 votes
1 answer
964 views

R limma alternatives in Python

The R package limma is ideal to perform differential expression analysis. Is there any limma alternative in Python? I'm trying ...
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1 vote
1 answer
60 views

Help with DIA-Mass Spectrometry data analysis with several conditions (limma?)

I am trying to learn how to analyse normalised DIA-MS data and I am struggling with it :// The original dataset I got is (6 conditions (2 samples each)) with 3 technical replicates (total: 36 sample ...
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1 answer
102 views

Toptable error, wont recognize condition

I am getting a rather strange error from topTable. When I run my code I get Error in fit$coefficients[, coef] : subscript out of bounds from topTable as if it is ...
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2 answers
126 views

Determining what RNAseq data is filtered on volcano plot

I am using RNA seq data to analyze genes via a volcano plot comparing differential gene expression of bacteria with and without antibiotic in R. After having created my plot, I am unsure why some of ...
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How is the t-statistic value calculated in GEO2R or Bioconductor?

I was trying to calculate t-statistics using Python's scipy and numpy as np- ...
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Troubles implementing LIMMA for paired samples (before/after treatment) comparaisons

i try to implement LIMMA for paired samples in order to compare gene expression before/after treatment. But...i'm not confident in my results since every single gene expression seems to be ...
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1 answer
673 views

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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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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3 votes
1 answer
1k views

Removing Batch Effect in Heatmaps after Differential Gene Expression Analysis

I'm working on a dataset in which the first replicate of each group is one batch and the second replicate is in a second batch. After checking the PCA plot and ...
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0 answers
160 views

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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2 answers
36 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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How can I get the estimate of lmFit function form limma package

This is the first time that I used lmfit function from the limma package I'm a little bit confused on how to interpret the result. is there a way to get the ...
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1 answer
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Can I use a regular liner regression model when I'm working with DNA methylation data?

I'm new on working with genetics data and I'm just wondering if I can use the lm function when I'm building my models or do I have to use ...
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2 votes
2 answers
2k views

TPM or rlog(CPM) for comparing expression?

I want to see the expression of a gene in a group of patient amongst the entire cohort using my RNA-Seq data. While I can do a differential expression analysis with limma or DESeq2, I want to see how ...
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1 answer
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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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1 answer
201 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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1 answer
96 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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2 votes
1 answer
90 views

Any way to filter out highly correlated genes with limma linear model?

I have Affymetrix gene level expression matrix (genes in the rows and sample ID on the columns), and I have annotation data of microarray experiment observation where sample ID in the rows and ...
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2 answers
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Any way to quantify the variation of genes that expressed in Affymetrix expression data?

am experimenting preprocessed Affymetrix microarrays expression data matrix (Affymetrix probe-sets in rows (32830 probesets), and RNA samples in columns (735 samples)) for my downstream analysis. Here ...
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1 vote
0 answers
112 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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2 votes
0 answers
55 views

How can `limma` be used to determine genes predictive of a given phenotype?

Using limma within Bioconductor here, which is a linear model package for microarray data and lmFit on gene expression data, I ...
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1 vote
1 answer
143 views

Voom transformation of RNA seq raw counts data

I have RNA seq data of raw gene counts that I want to transform for linear modelling. I am trying to voom transform, to do a weighted analysis. Data frame: prac_count_10 ...
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2 votes
1 answer
212 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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2 votes
1 answer
138 views

Significant gene set testing - limma

While I have used my title, it is more of a general statistics question regarding how these significance tests are carried out, these include decideTests and ...
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3 votes
0 answers
234 views

Differentially methylated position analysis in a related sample?

I'm trying to figure out how to do a DMP analysis (using minfi dmpFinder) on a related sample (if it's even possible). Right now the code (not written by me) is: ...
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2 votes
0 answers
448 views

Different results between paired t-test, paired t-test-like (empirical Bayesian), and mixed effect model

I am learning the R package limma using its user guide. I need help on understanding the difference between the paired t-test, paired t-test-like (empirical Bayesian) and the mixed effect model in the ...
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5 votes
2 answers
381 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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