Bioconductor Forum

it successfully for a qPCR array with 96 genes. I was wondering if someone knows about a couple of papers where limma is used to assess DEGs in a qPCR array and not on another microarray platform or RNA seq. I know it's perfectly

limma

updated 7.6 years ago • Daniel Moreno

this has lead to me towards another question as I attempted to extend such concepts to another experiment wherein the sample size in each group is different.  For example, here is a dataframe modified...dropping four or six samples seems more of a sacrifice.  I begin to think of experiments which (assuming repeate…

edgeR

updated 11.2 years ago • Charles Determan Jr

this forum, I still cannot figure it out what we should do if we have a variable considered as "Batch" and we want to use `lfcshrink`. In the vignette of [DESeq2][1] appears that the shrinkage estimator `apeglm` cannot handle...batch treated1 treated single-read a treated2 treated paired-end b treated3 treated paired-end c untreated1 untreated single...D…

BatchEffect lfcshrink RNA-seq DESeq2

updated 2.6 years ago • Eva

I have been following the paper ["Differential methylation analysis of reduced representation bisulfite sequencing experiments using edgeR"](https...I end up with so few reads. ```r ## The following code is similar to the one on the RRBS edgeR paper ## (...) Coverage <- yall$counts[, Methylation=="Me"] + yall$counts[, Methylation=="Un"] min.coverage <- 8 nObs <- ncol(Covera…

limma edgeR

updated 4.2 years ago • altintas.ali

Hi, I have **2545** and **1402** genes in **Oncology Biomarker Panel** and **Precision Immuno-oncology Panel** respectively. I have **719** common genes between two panels. I will need to merge raw read counts from these panels for differential expression (we know experimental condition, chemistry for samples from the same patients makes this reasonable). However, I thought to do differential …

cancer deseq2 r rnaseq

updated 7.1 years ago • Fereshteh

<div class="preformatted"> I have a table in this format that comes from the function vennCounts: c1 c2 c3 c4 Counts [1,] 0 0 0 0 26 [2,] 0 0 0 1 27 [3,] 0 0 1 0 4 [4,] 0 0 1 1 6 [5,] 0 1 0 0 5 [6,] 0 1 0...div class="preformatted"> I have a table in this format that comes from the function vennCounts: c1 c2 c3 c4 Counts…

updated 13.9 years ago • lcarvalh@btk.fi

000 columns for 1.1M BP positions (rows) . The read.table and read.csv options fail because of the table size (but works if I only import 100,000 rows at a time). I know that Bioconductor has many packages for annotating sequences...classes, skip=1, na.strings="", sep="\t", nrows=1112880) #Error: cannot allocate vector of size 8.5 Mb #In addition: Warning messages: #1: In scan(file, what, nma…

annotation importing

updated 11.3 years ago • k.askland

differentialy expressed gene using the **plotCounts** function. When i plot them i can still see the batch effect in the image generated.I then refereed to your post https://support.bioconductor.org/p/76099/. There it is mentioned...that it will model the batch effect in linear regression not remove them also in the following threads it is mentioned that we can use limma package...removebatcheffe…

deseq2 normalization

updated 5.9 years ago • dhwani.dholakia

I have a clear batch effect that's caused by sequencing paired-end vs. single-end on different days. I'd like to correct for this in the DESeq2...I have a clear batch effect that's caused by sequencing paired-end vs. single-end on different days. I'd like to correct for this in the DESeq2 analysis as suggested in the vignette ("If there is unwanted variation present in the data (e.g. batch effect…

deseq2 limma RUVSeq

updated 5.7 years ago • 94133

ChAMP pipeline. I understand that the first SVD is used to identify the correlations and correct batch effects. However, I don't understand how I can correct the effect of cell types as it doesn't appear in my csv file. What should

ChAMPdata ChAMP

updated 23 months ago • simleopold11

regarding limma, and its use with our data. I come from a programming background, with some math training (in the distant past); I am sure many crucial statistical issues elude me. I am looking for help. Our experiment seeks

Plasmodium falciparum probe limma Plasmodium falciparum probe limma

updated 19.5 years ago • Paul Shannon

I am doing a RNAseq analysis we have an option to use rlog or the vst transformations. In your paper "Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2" you have this statement: > while the...also effective at stabilizing variance, it does not > directly take into account differences in size factors; and in > datasets with large variatio…

rna-seq deseq2 warnings nbinomWaldTest transformation

updated 5.7 years ago • andreia

div class="preformatted">Draft paper submission deadline extended: BCBGC-09 The deadline for draft paper submission at the 2009 International Conference...the authors. The conference will be held during July 13-16 2009 in Orlando, FL, USA. We invite draft paper submissions. The conference will take place at the same time and venue where several other international conferences

updated 17.0 years ago • John Edward

preferable > because you can discount in the model other sources of variation. For > instance batch effects, other confounding variables/covariates > (smoking?, bmi?) and so on that are not age-related. Yes, this is also what...or decreased variability with increasing age should come from, but maybe I am missing something, my experience with building such models is very limi…

GO GO

updated 12.8 years ago • Simone

Sequence Analysis & Alignment * SNPs and Haplotyping * System Biology and Modelling *Paper Submission Details:* * Authors are invited to submit original research contributions or experience reports in English...The submitted manuscript should closely reflect the final paper as it will appear in the proceedings of BIRD'08. * Papers should not exceed 15 pages i…

Genetics Pathways Cancer microRNA Genetics Pathways Cancer microRNA

updated 18.2 years ago • Prof. Roland Wagner

used combat (available as an R script) from Evan Johnson and Cheng li with good success for removing batch effects. I can send you the script. I have edited it so that it work a little better with BioC. Aedin On Dec 8, 2007 10:59 AM, James...at="" yahoo.com=""> wrote: Dear List, I am now analyzing some Affy data with obvious batch effect with different dose level, one conceptual question …

affy DOSE affy DOSE

updated 18.2 years ago • Aedin Culhane

datasets for solving complex scientific questions * Provide bioinformatics consulting and software training to junior researchers ## Required qualifications * University degree or postgraduate training in bioinformatics...computational biology * Extensive experience in the computational analysis of NGS datasets * Familiarity with Linux environment, version control and compute...clusters…

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updated 8.3 years ago • sergisayolspuig

expressed genes. I put together a combined edgeR-SVA script to try and correct for the batch effect (shown below). I can clearly see the  batch effect variable in the output of `` svaseq$sv `` (the first surrogate variable...expressed genes in each model. So, my questions are: 1. Is it possible/valid to account for batch effect when doing a pairwise comparison between groups of samp…

edger rna-seq sva differential expression

updated 8.9 years ago • mpj5142

the vignette. I want to duplicate and RNASeq values and plots of an RNAseq exmaple in the quantro paper, before I apply quantro to my own data. I applied quantro to Bottomly et al.'s dataset (PMID:21455293) downloaded from recount...steps (eliminating all zero genes and running the rlogTransformation) which were mentioned in the paper. I got different quantro statistics and plots than in the…

quantro RNASeq

updated 5.8 years ago • raf4

Hello! I have RNA seq data and I need to use combat to remove the batch effects. Somehow when I run it, it isnt actually doing anything. ``` dds <- DESeqDataSetFromMatrix(countData=data, colData...metadata, design=~~Batch + dex, tidy = TRUE) dds <- DESeq(dds, betaPrior=TRUE) normalized_counts <- counts(dds, normal…

BatchEffect DESeq2 RNASeqData

updated 5.0 years ago • Emma

Dear fellow DEseq2 people, I have a question concerning the usage of DEseq after batch correction. I am looking at different samples from different timepoints and as we collected two samples at a different...time in a different lab, I wanted to correct for batch effect. However I am not sure for the design. Should we still include the batches as part of the design even when we do combatseq.…

DESeq2 BatchEffect

updated 3.0 years ago • johannesschoeneich

cover this, and is helpful), our lab ran a ChIP-seq for H2AZ in three separate sequencing and IP batches. This is clearly evident on PCA as the samples cluster along dim1 primarily based on their batch identity rather than...their genotype. From our experience, we would expect a composition bias, and profile plots around the TSS and centered peaks corroborate this where...this histone variant…

ChIPSeq csaw

updated 13 months ago • Luca

the block function in lmFit Targets SlideNumber Cy3 Cy5 Batch Bioreps 1 WTCL WTCR 1 1 2 CoffeeAL CoffeeAR 1 2 3 CoffeeCL CoffeeCR 1 3 4 WTBL WTBR 1 4 5 WTBR WTBL 1 4 6 CoffeeAR CoffeeAL...WTAR<- topTable(fit2, coef=2, adjust="BH") Is it sensible to make a coefficent for each of the batches in my design with my set of arrays? So three extra columns? I am unsure if I have enough inf…

updated 16.4 years ago • Katrina bell

Hello,   I have a question about a differential expression analysis that I am doing using Limma on RNA-seq data. Basically, I want find differentially expressed genes between tumour and normal samples (in men and women separately), using as covariates ethnicity and age. To do so, I defined a model like this one: design <- model.matrix(~ ethnicity + age + tissue\_type) …

Limma differential gene expression batch effect correction covariates

updated 7.7 years ago • abelsousa

Hi Yen Lin, I've read your posting from 11 Feb to the BioC list about the design of your experiment with two "batches" of mice. I've a similar experimental layout: mouse cell cultures treated with a drig at different...a principle component analysis of the cross-chip normalized probe sets I can clearly see that the batches cluster together. I've used affyPLM to shed some light into the experim…

Normalization Clustering probe affyPLM Normalization Clustering probe affyPLM

updated 22.0 years ago • Arne.Muller@aventis.com

generated by the rlog function from DESeq2: ```r dds <- makeExampleDESeqDataSet() dds$batch <- factor(rep(c("A","B"), each = 3)) rld <- rlog(dds) mod <- model.matrix(~ condition, colData(rld)) assay(rld) <- limma::removeBatchEffect...assay(rld), batch = rld$batch, design = mod) ``` Would it be correct to simply inverse the log2 transformatio…

DESeq2 limma

updated 4.6 years ago • jma1991

Dear all, about correcting the batch effects in **LIMMA and SVA,** 'd appreciate having your comments : assuming that we have a set of RNA-seq data (no treatment, + treatment...in many distinct BATCHES, would the linear model in LIMMA : **design <- model.matrix(~0 + BATCH + Treatment, data=RNA)** suffice to correct for batch effects

limma combat sva

updated 6.6 years ago • Bogdan

Post1-Pre1 `` `` Post2-Pre2 `` `` Pre2-Pre1 `` `` Post2-Post1 `` But taking into account batch effects (batch1 and batch2) and sex (male/female) . But I am not sure if including batch effect because Pre and Post samples...coming from the same individual fall in the same batch. This is my limma code so far: <pre> head(info) </pre> <table border="0" cellpadding="0" cellspaci…

limma paired samples complex-design

updated 10.1 years ago • aec

66 sample and want to employ the ARACNe method. The literature states that 100 is the minimum sample size for ARACNe but sample size testing was only done on microarray data. Given the more accurate assay (less noisy) in RNA seq

grndata GRN ARACNE WGCNA

updated 3.6 years ago • Harrsha

Dear edgeR support team, In the tutorial its mentioned that Batch effects can be removed or corrected by using the additive model formula. How can I visualise the removal of batch effect

edger batch effect

updated 11.1 years ago • candida.vaz

strains of *Candida albicans* across four time points and want to do WGCNA, but I only have TPM tables. The raw data exist (somewhere in our server...) but I'm unable to find them and just want to do WGCNA to have an idea of clusters...I can look at. I've been working off of TPM tables generated by a previous grad student and have done all my DGE analysis using edgeR, but the output of a DGEList …

RNASeq WGCNA ComparativeGenomics

updated 15 months ago • siobhan.dietz

I need some advice. I'm lookin at PCA plots of RNAseq data, and am understand whether my data has batch effects or not.  I performed alignment using STAR, and then obtained gene counts, and gene TPM values I used \`\`\`prcomp...drive.google.com/open?id=14buDDn8L11BHDgPzsCWXKJX0w70BlXp-)__ It seems that there could be a batch effect, but I'm not a 100% sure, since I'm doing this for the…

batch effect pca

updated 7.6 years ago • K

and tested for association between each PC and independent experimental variables like BCD batch (2 batches), Experiment batch (3 batches), Sentrix ID (9 chips), Sentrix position, cell components and sample groups using linear...regression. I find large amount of variation in my data due to BCD batch, Experiment batch and Sentrix ID even after normalization and I wish to adjust for these batch e…

ENmix combat.mc illumina450k combat sva

updated 9.7 years ago • DSP

div class="preformatted">how to set library size in DESeq? i know how to set library size in edgeR d <- DGEList(counts = d, lib.size = c(9893630,11055814,11207084,9663487,11455088...8140053), group = group)#??DGEList?? but donot know how to set library size in DESeq it seems i can set size factor sizeFactors(object) <- value or pData(cds)$sizeFactor <- value but how to se…

updated 14.1 years ago • wang peter

that I would like to analyze in `` limma `` has the following properties: * Seven separate experiments, which share a common chip and image-reading technology. * All RNA sources across experiments are biological replicates...I see two options to achieve this goal of ours: 1. A one-step approach where all arrays from every experiment are included in the `` limma `` analysis. In order to acco…

r limma limma design matrix

updated 10.5 years ago • mwesthues

div class="preformatted">Hi, How is it possible to judge whether there is any batch effect in two groups of Affymetrix .cel files ? I have got currently one Affybatch object by reading all the .cell files

updated 13.5 years ago • santana sarma

trying to figure out if there is a way to deal with technical replicates when coming from different batches of sequencing. I do not want to remove the batch effect from the data, I want to include it in the statistical model. So...Would I be able to apply this to my technical replicates knowing that they come from different batches? In this case, different batches mean the samples are run at a…

technicalReplicates BatchEffect limma RNASeq

updated 22 months ago • catv

responsible for the statistical analysis of high-throughput data from next generation sequencing experiments (RNA-Seq, ChIP-Seq, WGS, etc.) from bulk and single cells as a service to users from academia, hospitals, and industry...Support in statistical considerations of sample size and experimental design will be a regular discussion topic with customers. Internally, you will participate in the..…

job biostatistics bioinformatics data analysis core facility Job

updated 6.6 years ago • Michael Prummer

Hello, I’m working with cummeRbund in R 3.1.1 and I’m trying to get a different point size for my csVolcano plot. I’ve tried two approaches an neither see to work. after loading library and the data as <pre> library...height:1.6">v <- csVolcano(genes(cuff), “EP”, “PC”, showSignificant=TRUE, alpha=0.05, mapping=aes(size=5))</span></pre> <span style="line-height:…

cummerbund ggplot2 size r volcanoplot

updated 11.2 years ago • ianmisner17

a question about the function "ComBat" in the package "sva". I am using the ComBat to remove the batch effect in methylation data. For the test data, the sample information is: sample covariate batch 101 1 A 102 1 A 103 1 B 104...1 B 201 2 A 202 2 A 203 2 C 204 2 C There are 3 batches and 1 covariate with two el…

updated 11.6 years ago • Guest User

I have used `` limma `` trend to find DE genes between 3 groups (including batch effect in design). Now I would like to visualize the genes, and used `` removeBatchEffect() `` to get corrected logCPM values...for each sample. `` logCPMc <- removeBatchEffect(logCPM, batch=targets$Batch, design=design_4_batch) `` Now I want to use the average of each group, and with `` aveLogCPM() `` from …

edger limma cpm

updated 7.4 years ago • b.nota

with Long lab and HIP Core researchers to design, implement, and analyze new immunological experiments. - Evaluate emerging computational methods and tools for cytometry data processing and analysis. - Recommend...extend, and implement best-of-class methods (i) to integrate cytometry data from multiple batches and projects, (ii) to integrate cytometry data from different p…

flowWorkspace CytoGLMM FlowSOM

updated 4.3 years ago • hbolouri

div class="preformatted"> I have a question regarding Batch and Covariates in ComBat. I have an Illumina expression dataset of control versus treatment, at 3 different time points...and by default samples cluster together based on the RNA extraction date therefore I have a strong batch effect. Should I use RNA extraction date as Batch and both the biological replicate groups and Time_x_Treatm…

updated 13.1 years ago • Guest User

Hello everyone,  I am using SVA for batch effect removal. I would like to know if it is possible to find out that batch effect removal procedure has actually worked...and now there is no batch effect? or simply to visualize data before and after batch effect removal and see how data has transformed? Thank you

combat sva batch effect comba

updated 9.3 years ago • amoltej

In trying to combine 67 Affy u133a and 67 Affy u133a_2 cel files I am able to form the initial affy batches using read.affybatch() but get a memory allocation error (below) when I try to combine them with the 'combineAffyBatch...function Error: cannot allocate vector of size 280,470 Kb Attempts to increase memory size within 'R' are only partly successful. Attempting to increase the memory to..…

affy affy

updated 19.8 years ago • Carleton Garrett

<div class="preformatted">Dear all, We have 63 arrays that are either diseased or normal. We wish to select genes adjusting for two covariates : gender (male/female) and experimental batch (one/two/three). >From biological knowledge, we expect the batch effect to be significant but we wish to quantify it numerically. Here is a way that we tried and the problems we faced. We searche…

Cancer limma Cancer limma

updated 19.9 years ago • Adaikalavan Ramasamy

Hi, I'm currently exploring the functions in DESeq2 and try to replicate some findings of DOI: 10.1126/science.aan3456. In this paper, they test several models to identify clusters of differentially expressed genes between 3 species (H[uman]/C[himp]/M[acaque...the functions in DESeq2 and try to replicate some findings of DOI: 10.1126/science.aan3456. In this paper, they test several models to …

deseq2 LRT

updated 6.0 years ago • bio_inc

variable (Control vs Patient), a developmental timepoint variable (Diff vs Undiff) and 2 unequal batches. I want to compare Patient vs Control in both Undiff and Diff states, but I must remove batch effects (MDS plot showed...batch 1 and batch 2 clusters). See below for the factors I created and the layout of the different groups. Disease <- rep(factor...c("Ctrl", "Patient")), each…

egdeR design matrix batch effect

updated 6.3 years ago • ahsindoomilup

<div class="preformatted">I've been testing out the pamr package and have read the paper "Diagnosis of multiple cancer types by shrunken centroids of gene expression" by Tibshirani, et. al. In the paper it mentions using 10-fold cross-validation. The pamr.cv function defaults to setting nfold to the smallest class size, in this case 8. Using the khan.txt tutorial data I've tried setting nfo…

Cancer pamr Cancer pamr

updated 18.0 years ago • Patricia Greninger

div class="preformatted">Dear Dietmar, Thank you very much for the feedback of the paper and the package! Where did you download the package and what is the package version? It could be that the dataset was not...dfernandez045 at="" ikasle.ehu.es=""> wrote: > Dear Dr. Zhu, > > I have just read the paper you and your colleagues have written > "ChIPPeakAnno: a B…

Transcription ChIPSeq Annotation chipseq genomes Transcription ChIPSeq Annotation chipseq

updated 15.7 years ago • Julie Zhu

BioAge Labs is a clinical-stage venture-backed biotechnology company headquartered in Richmond, CA. We’ve built a systems biology platform to map out the key molecular pathways that impact healthy human aging, based on proprietary human aging cohorts that have blood samples collected up to 45 years ago with participant-omics data that is tied to detailed medical follow-up records over their lifes…

BioAge ComputationalBiology JobPosting

updated 4.8 years ago • jamie

attachment. Now re-sending without attachment: I was having a problem with ComBat when using large batch sizes and the non-parametric parameter estimation. Almost all probe intensity values after ComBat correction would...below: # Monte Carlo integration function to find the nonparametric adjustments for a particular batch int.eprior <- function(sdat,g.hat,d.hat){ g.star <- d…

probe probe

updated 7.9 years ago • Jason Stein

What was the very first paper to describe "limma-trend"? By "limma-trend" I mean the one described in Law (2014), but I am not sure that was the first time. Thanks

limma

updated 10.5 years ago • Nik Tuzov

I am using DESeq2 to investigate changing microbial taxa after incubation experiments. I am comparing my Preaddition community to 5 post-incubation communities (different nutrient additions &amp...change. I have approximately 49,000 OTUs (Operational Taxonomic Units) in the original count table which includes all of the incubations, and I am individually investigating the different incubation…

deseq2

updated 9.0 years ago • tborchardt2

participating and presenting his/her work in this workshop may do it by sending a contribution. The papers should be written in English. There are three possibilities: 1- Long papers: A maximum length of 8 pages. They must consist...progress of their research, in order to obtain feedback from a panel of experts. The length of these papers should be no longer than 8 pages. - Papers Format Papers…

updated 17.2 years ago • Nora Muda

mcols(object)$fullBetaConv. Use larger maxit argument with nbinomLRT" all the time. but I created batch as a factor based on what I see in the data. I tried with and without it. ```r Batch = c("1", "2", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3", "3") ``` ```r dds=DESeqDataSetFromMatrix...countData = countData, colData = colDa…

batch svaseq plotPCA fullBetaConv

updated 3.7 years ago • kin_ket

wrote: > >Dear Sohail, > > > >Well, there are lots of ways to generate such a table. Perhaps the simplest is > > > > fitsel <- fit2[sel.dif, ] > > as.data.frame( fitsel ) > > > >Best wishes > >Gordon...gt; > >From: "Khan, Sohail" <khan ...="" …

limma limma

updated 15.1 years ago • viritha kaza

is: paired samples before and after treatment, and I would also like to take into account batch effect. I am having problems with the batch effect, by reading through mails in this mailing list, I've gotten...to do: Create example date, patient (5 patients, 2 samples), treatment (A or B), and batch (three batches, 1-3) <pre> >…

limma

updated 10.7 years ago • John Burger

Island, and Penglai Pavilion Scenic Area. Seafood and fruits are plentiful in Yantai. Selected best papers will appear in SCI-indexed journal(s). All papers in conference proceedings will be indexed by both EI Compendex and...i.e., China), foreign experts are encouraged to propose invited sessions. The first author of each paper in an invited session must not be affiliated with an organization i…

updated 16.3 years ago • ICNC'10-FSKD'10

Hi, I have 26 samples from 12 patients sequenced in two batches. 6 of these are "normal" samples while the other 20 are tumors. What I have done before is simply compare the tumors against...the normals, and include the batch variable as follows: design.file$batch <- as.factor(design.file$batch) dds <- DESeqDataSetFromMatrix(countData...read_counts, colData = design.file…

deseq2

updated 6.6 years ago • sxv