Bioconductor Forum

data to analyze with 3 factors: Genotype (wt, mut), Treatment (C, T), and Organ (s, r), each in 3 replicates. I found the guide for limma, I am just confused about how to "makeContrasts" to get all levels of main and interaction

interactions 3factors limma Microarray

updated 4.9 years ago • Dan

RNAs are isolated from 42 patients. could you tell me how can I set the design matrix, there is no replicates and it is only one condition which is compared over time (i.e there is no treatment factor). Thank you for your help

Microarray limma Microarray limma

updated 16.6 years ago • claire pujoll

to create (or have) a similar set of files? Generally, we have only 2 fastQ files - R1 and R2. Are replicates essentials? Can I skip the normalization step? Also, what about coupling set. How should I go about it

bioconductor MEDIPS

updated 9.2 years ago • Vijay Lakhujani

div class="preformatted">Hi all, I want to ask a question about using DEXSeq. I have two technical replicates for one sample and I need to merge them to get the exon counts. I've mapped them and get the sam files, can I just merge

DEXSeq DEXSeq

updated 12.6 years ago • chunjiang he

<div class="preformatted">Dear All, I have carried out an RNAseq experiment with 4 conditions, 2 biological replicates of each. In the moment, I am interested in how my conditions differ in terms of expression of a subset of 36 genes. The...preformatted">Dear All, I have carried out an RNAseq experiment with 4 conditions, 2 biological replicates of each. In the moment, I am interested …

RNASeq RNASeq

updated 13.9 years ago • vladimir mashanov

trying to use edgeR for analysis of gene expression across 4 RNASeq samples(different stations) (3 replicates each.  The information I would like to have, is the expression (fold change) for each gene across all 4 stations

edger differential gene expression GLM Tutorial

updated 9.8 years ago • wendy_11

FDR 5% using cufflinks whereas just found 50 upregulated genes at 10% using DESeq. I dont have any replicates. > Also does DESeq overcome length bias, a general problem in RNA seq data analysis? Regards Aniket [[alternative

DESeq DESeq

updated 15.4 years ago • Aniket Vatsya

I keep getting this error with autoplot (or plotIdeogram) in ggbio. <pre> Error in rep(startY, each = length(yy)) :    attempt to replicate an object of type 'language'</pre> Wondering if it has to do with the ggplot2 (3.1.0) as suggested by this [issue](https://github.com...plotIdeogram) in ggbio. <pre> Error in rep(startY, each = length(yy)) : …

ggbio ggplot2

updated 7.1 years ago • t.fadason

I am doing edgeR glmLRT() test because I don't have any replicates. I have three groups and I made the design as: design <- model.matrix(~0+group+group:plant+treatment:group,data

edger design matrix glmlrt

updated 9.0 years ago • simarsidhu25

and perform differential methylation analysis between groups in Lumi. For example data contains replicates and three goups as non-methylated, methylated, hemimethylated. I wanto group samples according to their methylation

lumi lumi

updated 14.2 years ago • rajasereddy

and also FPKM values for these genes. To my knowledge, I need to plot Z-score values from the 3 replicates per sample for each gene (rows) but I dont know how to get Z-scores. Should I calculate the Z-scores first and then read

normalization pheatmap

updated 5.7 years ago • Hamidreza Hashemi

<div class="preformatted">Hi when I replicate the example for exon array data in the GenomeGraphs vignette and set displayProbesets to TRUE then the names of...div class="preformatted">Hi when I replicate the example for exon array data in the GenomeGraphs vignette and set displayProbesets to TRUE then the names

Cancer graph Cancer graph

updated 16.7 years ago • Max Kauer

I'm curious why that would be (sorry, stats isn't my strong suit). For example (each condition has 3 replicates): Condition 1 normalized counts: 18536.58274, 20600.98049, 22578.73105 Condition 2 normalized counts: 12705.48353

deseq2

updated 8.3 years ago • Mike

edgeR, deseq) that calculate the list of differentially expressed genes in 2 conditions (with replicates) from raw counts. But I do not know what is wrong with the following simple approach (and whether other people have

updated 11.3 years ago • Son Pham

gene expression analysis by myself instead of using wald test the package provided as I don't have replicates in each comparison group. so, there is no statistical calculation but only fold change between two samples. I am

deseq2

updated 10.8 years ago • Xiaokuan Wei

Hi, For few of my plant samples with no biological replicates I am using edgeR. If I use bcvvalue 0.1, I am getting >10k significantly differential expressed transcripts...Hi, For few of my plant samples with no biological replicates I am using edgeR. If I use bcvvalue 0.1, I am getting >10k significantly differential expressed transcripts out

edger

updated 8.9 years ago • manoharankumar01

<div class="preformatted">Hi, My problem is as follows: I have a bunch of subjects with a medical condition, and a bunch of normals, and I am looking for differences between the two groups. However, I have several arrays per subject (biological replicates), and so would like to include subject variability into the model. If this was a usual single-response problem, I might...for dif…

updated 20.2 years ago • rob foxall IFR

A B IP 2 2 IN 2 2 The number 2 indicates 2 biological replicates for each cell. When DiffBind calls DESeq2, what contrast does it use internally to calculate LFC and FDR between

diffbind

updated 6.1 years ago • liruiradiant

meaning I should control for a batch effect for the model (original nest, in the model as Replicate). I approached the analysis as follows: ```r dds <- DESeqDataSetFromMatrix(countData = tempdf, colData = sfile, design...Replicate + Treatment + Time + Treatment:Time) dds <- DESeq(dds) ``` Using contrasts, I wa…

GeneExpression DESeq2 TimeCourse

updated 14 months ago • Jason

<div class="preformatted">hi to all bioconductor users! our goal is to find differentially expressed genes in cell line and patient probes with and without a given treatment. so usually we have two chips (we have not enough money to do replicates, i think that's fairly common with affymetrix ;) ), one chip has RNA with treatment, the other one without. i currently try to find out what norm…

Normalization affy GLAD Normalization affy GLAD

updated 21.5 years ago • Dipl.-Ing. Johannes Rainer

is not the same in the two samples. If you have two different experimental conditions, with replicates for each condition, DESeq tests, whether, for a given gene, the change in expression strength between the two conditions...is large as compared to the variation within each replicate group." Current language on the Cuffdiff site suggests that the current version of that program tests for whet…

Sequencing Bayesian Cancer DESeq Sequencing Bayesian Cancer DESeq

updated 13.9 years ago • Richard Friedman

<div class="preformatted"> Hi I am writing as I am trying to analyse NGS data from public data (GEO) specifically datasets such as one sample per time point. The raw (somewhat processed data) is 3 samples at different time points where ???The read count at exon, splice-junction, transcript and gene levels were summarized and normalized to relative abundance in Fragments Per Kilobase of ex…

DEGseq DEGseq

updated 13.3 years ago • Guest User

data for three antibodies. I am just interested in data for one antibody, which has two biological replicates. I guess it's better to include only the two samples I am interested in normalization (the step preprocess). Any comments...4) In the computeRunningMedians step, because I have two replicates and want to combine them, I should use combineReplicates=TRUE. Am I correct? Thanks in advance! …

Normalization Ringo Normalization Ringo

updated 15.3 years ago • Yong Li

and is intended to deal with more subtle issues. Normalization, as edgeR does it, does not require replicates. Best wishes Gordon > Date: Fri, 04 Feb 2011 11:28:15 +0100 > From: Jens Georg <jens.georg at="" biologie.uni-freiburg.de...normalization with edgeR and do you think a reliable basic normalization > is possible without replicates? > > Thank you for…

Sequencing RNASeq Normalization edgeR Sequencing RNASeq Normalization edgeR

updated 14.9 years ago • Gordon Smyth

Hi all, I am analyzing an rna-seq dataset (treated vs control) with three replicates in each. After performing differential expression with DESeq2, there are way too many differentially expressed...5) before running DESeq, but still I have many DEGs. PCA results show nice clustering of samples and replicates so no problem there.    Is there any thing I am missing ? Thanks. …

deseq2 rna-seq

updated 9.4 years ago • anand_mt

div class="preformatted">I have a reasonable RNASeq data set of 10 biological replicates of a control group versus 10 biological replicates experimental I've gone through the edgeR workflow, and get

RNASeq edgeR RNASeq edgeR

updated 13.7 years ago • Simon Melov

where Gordon suggested using a simple excel formula to achieve fold changes when you dont have replicates *lib.size1 <- sum(y1)* >>* lib.size2 <- sum(y2)* >>* logFC <- log2((y1+0.5)/(lib.size1+0.5)/(y2+0.5)*(lib.size2+0.5))* * * Is this...moment to see if this analysis works and obviously when I come to more complicated public data with r…

Transcription DEGseq Transcription DEGseq

updated 13.3 years ago • Jill Pleasance

According to the experimenters they require 10 ug per chip) So it is not possible to use biological replicate chips for each individual mice. Now the issue is whether to perhaps pool the RNA in each group and carry out analysis...on technical replicates from the pooled samples. As I understand it pooling may reduce the precision, with the risk that one or few samples

updated 22.3 years ago • Wiesner Vos

preformatted"> Greetings friends! I seek help with data that I have : 3 time points, 3 genotypes, 3 replicates for each of these = 27 libraries The goal is to find genes that have different time expression profiles amongst...removing genes that have low expression (count) levels removing genes that have high variance across replicates removing genes that have low variance across time (consti…

Normalization edgeR Normalization edgeR

updated 13.3 years ago • Guest User

Hi, I have ChIP peaksets for three different factors in two different treatment groups. For each I have several replicates. I have generated a consensus peak set using: consensus<-dba.peakset(dba, consensus=c(DBA_FACTOR,DBA_CONDITION,DBA_TREATMENT), minOverlap=0.99) Now I would like to plot the overlap between the consensus for FACTOR1 and TREATMENT1 and the individual replica…

DiffBind consensus peaks venn

updated 6.3 years ago • neumann.sylvia

I am using limma to analyse a 2-colour microarray experiment. There are 2 >>treatments and 4 replicates in each of these groups. Each replicate is >>paired to a replicate in the otehr treatment group. Each sample was

Microarray limma Microarray limma

updated 21.6 years ago • Naomi Altman

Hello everyone! I've run the nf-core Cut&Run pipeline on five samples, each with three replicates, using IgG as a control and an E. coli spike-in for normalization. The pipeline normalization was set to spike-in...Hello everyone! I've run the nf-core Cut&Run pipeline on five samples, each with three replicates, using IgG as a control and an E. coli spike-in for normalization…

Normalization DiffBind

updated 14 months ago • kim

Dear community, I was using DESeq2 to analyse a small set of data (7 conditions x 4 replicates = 28 samples). I only keep genes that have more than 10 counts in more than 2 samples. Now I got a very strange dispersion

DESeq2

updated 3.4 years ago • Yiqi

counts): ![Mock matrix][1] I'm assuming that each species in such matrix could be considered a replicate within each group, that gene families would be the equivalent to loci, and counts are absolute. Any suggestion/advice

edgeR

updated 3.7 years ago • eliza_3176

Q1: Is there a way to compare and contrast 6 time-points, considering each has 2 biological replicates. Q2 If the answer to Q1 is no, when I compare A to B and separately A to C , why do I get different counts for A in the binding

diffbind DiffBind

updated 4.1 years ago • akankshabafna

to do differential gene expression analysis on my mRNA seq data. I just have two group, each of 5 replicates, that I want to compare. Out of curiosity I performed both the classic EdgeR (exact test) and the GLM (glmQlFtest) approaches

DifferentialExpression edgeR

updated 2.1 years ago • Jurgen

nematodes. I have differential expression data (15k genes) in FEMALEvsMALE conditions, each with two replicates (drf1, drf2) for FEMALE and (drm1, drm2) for MALE. I have been exploring the WGCNA package in R to obtain gene co-expression

RNA-Seq WGCNA Worms

updated 2.1 years ago • davca

Hi Michael Love, I am pretty new in this. I have "wt" and "ko" RNA-seq samples, each three replicates for differential gene expression analysis. sample description as follows: wt group = 2 males and 1 female ko group

DESeq2

updated 2.4 years ago • Mahendra

got errors that kept me from including it in the end. I believe the error stemmed from a lack of replication within each batch. Is it possible to account for a batch effect using this design? ![enter image description here

DESeq2

updated 3.6 years ago • Stephan

seq analyisis using R. I Am usig DESeq 2 for my analysis. I have four different Samples with three replicates each and I want to compare all of them with control plk. I tried using contrast but i can only compare two factors

DESeq2

updated 3.6 years ago • Manmohit

Help me In deseq2 design I have factors sex, 2species, 5stages I want to know differential expressed genes between male and female of one species and And then from this extract I want to compare male of this species to 2nd species male In short Regress out female effect from each stage and other species male I designed like Design = ~ Replicate + sex + stage + species An…

DESeq2 genes Differential expressed

updated 2.7 years ago • Syed Zaheer

how to add extra data, so I wondered if it is possible to change existing data for  Tissue or Replicate?      Kind Regards Lauma&nbsp

diffbind. change of metadata

updated 7.6 years ago • Lauma R

I am wondering how best to incorporate the longitudinal data.  Would this be best used as a replicate?  Or better to model each patient as an explanatory variable? Thanks Eric

limma microarray

updated 11.1 years ago • Eric Zollars

For differential gene expression analysis of three treatments, each of which has three replicates, we find the results are different when we used fitType="local" versus fitType="parametric" in DESeq2.  For the

deseq2

updated 8.7 years ago • liangc.mu

count, k) will initialize the EM algorithm using a kmeans heuristic for selecting the starting point. Replicate runs on the same data can yield stark differences in the result. I have a dataset in which it seems that naively chosen

updated 11.5 years ago • Charles Berry

below. library(MSnbase) library(pRolocdata) data(dunkley2006) dun <- split(dunkley2006, "replicate") exprs(dun\[\[1\]\])\[1,1\] <- 1 __Error in \`\[\[<-\`(\`\*tmp\*\`, 1, value = <S4 object of class "MSnSet">) : __ __  \[\[<- defined for objects of type

msnbase

updated 7.8 years ago • kamal.fartiyal84

my time course (0,1,3,5 and 7 days) in 2 different mice genotype (Wild type and Knockout). I have a replicate in all the time points. I want to understand what genes (microarray) are DE during my time course in the wild type as

limma microarray

updated 6.7 years ago • ijvechetti

Gene expression analysis.  I have read counts data on 50 individuals in three biological replicates. I would like to filter out lowly expressed genes. Is there a threshold to define express genes?   I was thinking

edger rnaseq

updated 8.8 years ago • myprogramming2016

Dear community members, I am currently working with the microbiome analysis of a plant using the RNASeq data. I have 3 replication with 2 treatment data. I have normalized count data of each microbe in the above data sets. I was wondering if I could...I am currently working with the microbiome analysis of a plant using the RNASeq data. I have 3 replication with 2 treatment data. I have n…

edger

updated 5.4 years ago • megha.hs28

Hi, I need help for using one way anova for microarray data. In 20 files, I have 5 treatment and 4 replicates for each treatment. I have the following code: . . . treatment <- gl(5,4,20, label=c("T1","T2","T3","T4","T5")) # define ANOVA function aof

Microarray Microarray

updated 13.1 years ago • listas de consultas

expression analysis for a time course experiment. I have six time points. At each time point three replicate RNA data were collected. Samples within and between each time point are independent. I am interested in checking

deseq2

updated 8.8 years ago • yohannesafew

<div class="preformatted"> Hi there, I am trying to analyze chip-chip data from Nimblegen 385k arrays for histone methylated regions. I have replicates from both wild type and knock out cells with separate inputs (controls) for the KO and WT. That makes a total of 12 arrays...trying to analyze chip-chip data from Nimblegen 385k arrays for histone methylated regions. I have replicates from …

updated 15.9 years ago • fire1976 wyoming

this? Secondly, is there a bioconductor function that identifies outlier spot intensities among replicate arrays? Appreciate your response. Anjan -- =================================== anjan purkayastha, phd. research associate fas center for systems biology

Normalization Normalization

updated 15.3 years ago • ANJAN PURKAYASTHA

I am comparing the gene expression of a bacterium grown in two conditions (~ 10 replicates each). I am not interested in the mean gene expression differences between the two, rather I am interested in variability

deseq2 variation

updated 5.6 years ago • dannyc

OF REPLICATED NonControl Probes reps 7 8 10 15 30 630 630 414 3094 6 ------------------------------------------------------ Replication at Probe level- MEDIAN CV </pre>   Each gene has 30 probes (one or more than 2 different probes). As you can see...In summary: Each gene has one or more probes and a total of 30. The distri…

limma mirna microarray

updated 9.9 years ago • roser.navarro

Hello Bioconductor, I'm am trying to understand how the functions of group_by and reduce in plyranges package work. My purpose is to create a Granges object that has all the genomic ranges from BAM files and reduce them, to find the common / union in all samples. example: # toy samples taken/modified from BiocWorkshops/fluent-genomic-data-analysis-with-plyranges set.seed(42)…

plyranges granges

updated 6.2 years ago • Konstantinos Yeles

Transcript quantification was done using Salmon with `--numGibbsSamples=50` to generate Gibbs replicates. I used the `catchSalmon()` function to import both the quant.sf files and the inferential replicates. edgeR utilizes

RNASeq edgeR LongRead Transcript

updated 5 months ago • mohammedtoufiq91

have been hybridised to an array designed for the model plant. There are only 2 chips - 2 biological replicates with dye swap. I normalised the data with limma as below - RG.nb <- backgroundCorrect(RG, method="none") MA.nb <- normalizeWithinArrays...show a good correlation? Also in more general terms I've been looking at the correlations between replicates (R and G values, not rati…

limma limma

updated 21.5 years ago • Matthew Hannah

see below)? Set up: I have a control and three dose groups (low, medium, high) each with five replicates. Within each treatment group, samples came from one of twelve different mesocosms, with each mesocosm representing...nbsp;After reading the EdgeR manual, I tried blocking for mesocosm, but because I don't have replicates for each mesocosm, I got an error message due to my design matrix not ha…

edger rnaseq

updated 6.1 years ago • emb13

a samplesheet.csv describing my samples and it looks like this: SampleID,Tissue,Factor,Condition,Replicate,bamReads,bamControl,Peaks,P eakCaller meio.1,meiocytes,H3K4me3,N,1,M_meiocytes_H3K4me3.bam,InM_input_meiocyt...gt;H3K4.B73 2 Samples, 38870 sites in matrix (45304 total): ID Tissue Factor Condition Replicate Peak.caller Intervals 1 meio.1 meiocytes H3K4me3 N 1 …

DiffBind DiffBind

updated 12.3 years ago • Anitha Sundararajan