Bioconductor Forum

in RNA-Seq data using Ballgown? I have got RNA-seq data for different batches of sequencing for the replicates of the sample. I followed new tuxedo pipeline for my preliminary analysis of the data and found that there were

differential expression analysis ballgown r batch effects total number of reads

updated 7.3 years ago • amy16

of microarray. I have about 12 time points for a yeast data, and each time point only has two replicates. I am wondering if I can use quadratic model to perform the analysis. If I used higher order model, will overfitting

Microarray Yeast Microarray Yeast

updated 15.4 years ago • Xiaokuan Wei

Hello, I have 2 samples (3 replicates of each) of MeDIP, and I have questions regarding their analysis with the MEDIPSpackage: 1. I have one input DNA for

MEDIPS input

updated 9.5 years ago • GFM

<div class="preformatted"> Hi there, I am trying to use LIMMA to analyze gene expression data from an experiment which has dose response but only one replicate at each dose. I tried to fit a linear model using lmfit(). I used the doses as continuous variable. I do the ebayes fit and...to use LIMMA to analyze gene expression data from an experiment which has dose response but only one replic…

affy limma DOSE affy limma DOSE

updated 15.2 years ago • fire1976 wyoming

I have a set of Mouse RNASeq data for various points of a differentiation time series, with replicates. A known fraction of the cells in each sample are Mouse Embryonic Fibroblasts (MEFs), that fraction varies quite

deseq2 svaseq

updated 8.4 years ago • gpalidwor

primary vs. recurrent), so a total of 22 samples. I am unsure of how to specify the groups and replicates parameters in the "pairedData" class. There is no other grouping parameter. They are all cases. thanks alice -- output

baySeq baySeq

updated 12.3 years ago • Guest User

Each media condition was paired with an acetaminophen (APAP) treatment condition for each biological replicate: ?- no exposure (0 mM APAP), the control ?- exposure to 5 mM APAP ?- exposure to 10 mM APAP Expression levels were measured for...each biological replicate-medium combination using the two-channel Agilent 011868 Rat Oligo Microarray G4130A ([GPL890][4]). All gene expression...Channel 2…

Microarray Organism limma oligo Microarray Organism limma oligo

updated 14.5 years ago • Chris Lasher

in order to analyze a large number of Affymetrix microarrays. The experimental setup includes 2-3 replicates, 2-4 different time points and 1-5 different dosage levels. So a typical targets file could look like the following...nbsp;                 repli…

limma design and contrast matrix design matrix limma design matrix

updated 10.5 years ago • Moritz E. Beber

than DESeq at downweighting tags with extreme variances, or that this has to do with the number of replicates. While extreme cases like the example that Laurant mentions may need special intervention, edgeR was specifically...designed to downweight highly variable tags, and this is just as effective with few replicates as for many. Let's simulate a dataset with Laurant's tag as the first one: …

RNASeq qPCR edgeR DESeq RNASeq qPCR edgeR DESeq

updated 14.8 years ago • Gordon Smyth

between groups when no filtering was conducted. RNA-Seq experiment details: 24 biological replicates averaging 10 million mapped reads per replicate. 12 samples belong to group one and 12 samples belong to group...On perhaps a related note, should I alter my code because I have a fairly large number of biological replicates. Below is the entire code I am running. Many thanks for your advice! …

edgeR edgeR

updated 12.9 years ago • Mark Christie

<div class="preformatted">Hello mail-list Experiment This microarray experiment was conducted to study gene expression in bacteria at different growth conditions. There are 12 different conditions: > unique(conditions) [1] "gr46" "NaCl" "Glycerol" "HCl" "NaOH" "CH3COOH" [7] "EtOH" "gr15" "PK" "BC7ppm" "BC9ppm" "EtBr" The one called PK is the po…

Microarray Microarray

updated 21.0 years ago • Ingunn Berget

gt;I have a dataset which is pretty much IDENTICAL to the swirl dataset: > >Experiment 1 - two replicate arrays with a dye swap: > >TreatedCy5 vs UntreatedCy3 >UntreatedCy5 vs TreatedCy3 > >Experiment 2 - two replicate

GO limma Bioconductor

updated 3 months ago • Gordon Smyth

Dear all, I am attempting to perform DESeq2 analysis (using the Geneious plugin) for targeted RNA-seq of several hundred genes. Only the target set of genes is sequenced, as the primer that I use for first strand cDNA synthesis is specific to the target set of genes. I have three replicates for each condition. Based on the biology of the system that I am working in, I know with a high degree …

DifferentialExpression DESeq2

updated 4.0 years ago • dadca596

<div class="preformatted">Hi! I'm writing with a few questions about applying ComBat (sva package) to a set of ~50 samples run on the the Illumina Infinium HumanMethylation450 BeadChip array (~450,000 DNA methylation data points). There is a large amount of variation in my data due to both the batch the samples were run in (3 different batches), in addition to the position they were locat…

GO Category sva GO Category sva

updated 11.9 years ago • Magda Price

data to see if there is differential expression between experiment and control. I have 3 biological replicates of each and the experimental set-up would slightly favor a "_paired_" statistical approach. Nanostring nCounter...Another paper describes an FDR permutation approach, but they don't seem to have any biological replicates, but 32 control experiments and 10 control genes (Amit et al., 200…

RNASeq Normalization Preprocessing DESeq RNASeq Normalization Preprocessing DESeq

updated 14.5 years ago • Vanessa Vermeirssen

2], is notorious for assuming that the various libraries in the dataset can be treated as technical replicates; i.e, most genes are not differentially expressed between conditions. When analysing a cell differentiation...whether the VST method implemented in DESeq assumes that the samples can be treated as technical replicates? </p> <p style="box-sizing: border-box; margin: 0px…

deseq deseq2 vst

updated 8.7 years ago • jiab

<div class="preformatted">Hi, I guess the simple question is would you expect or have you seen a 'standard' distribution of p-values for a treated-untreated comparison (3 reps) after the eBayes procedure in Limma? Expressed in my usual 'comprehensive' style ;-) Following on from a previous thread I've started to look more into the p.value distributions to get an idea of false +ve and -ve …

limma limma

updated 21.3 years ago • Matthew Hannah

clones. Each of these clones is fed with two different diets (treatment).We have two biological replicates for each condition. Hence, we have a total of 12 RNAseq samples (3 different clones x two diet treatment x 2 replicates

RNASeq DESeq RNASeq DESeq

updated 13.2 years ago • Akiko Sugio

pre> > papivoc[1:6, 1:6] Names Sample1 Sample2 Sample3 Sample4 Sample5 1 Replicates healthy healthy healthy healthy healthy 2 C19757 2.1927358 1.6329086 2.4302252 0.5544893 <NA> 3 C00042...characters <pre> > str(papivoc) 'data.frame': 21 obs. of 81 variables: $ Names : chr "Replicates" "C19757" "C00042" …

papi metabolomics pathway analysis

updated 8.8 years ago • hp71727

do significance testing, and I'm not sure we can test for significant changes if we simply lump the replicates of cell-specific mRNA of a time point together and compare to the lump of replicates of Total-mRNA of the same time

deseq2 rnaseq ribotag trap

updated 6.0 years ago • mdfors01

hello, I am working on DEseQ2 to find the differentially expressed when i compare the control (3 replicates) Vs. 2 treated samples (3 replicates each, and 6 in total for treated condition) . I am mainly interested in the genes

rnaseq degseq

updated 8.2 years ago • laksharikrish

colData = exp_design, design = ~ Replicate + Treatment) ``` After that, I'd like to extract the counts from the normalized dds arranged by one of the samples, concretely

DESeq2 Transcriptomics

updated 21 months ago • Anita

Hi- I am new to analyzing Illumina microarray data. We ran 6 case and 6 control samples (no replicates) on 1 Illumina HT-12 v3 Expression BeadChip. I would like to analyze our data using lumi. I was planning on doing the

Microarray Normalization lumi Microarray Normalization lumi

updated 16.8 years ago • Christina Markunas

tried to obtain the same information with GO.db and rols, with no success. Specifically, I tried to replicate the use cases in the above link.  I tried to retrieve all the terms from GO.db and then checked all the GOXXPARENTS

gene ontology rols

updated 11.3 years ago • Michele

improves support for models with covariance structures: random effects models for technical replicates, mixed models for log-intensity analysis of two color data, duplicate spots etc. Lots of updates to help files. pdf

limma limma

updated 21.4 years ago • madman@jimmy.harvard.edu

I have used the normalizeBatch() function of the cydar package to apply batch correction to a mass cytometry dataset, based on an identical technical replicate present within each batch. The cydar package manual describes how to convert the output of normalizeBatch() to a...cydar package to apply batch correction to a mass cytometry dataset, based on an identical technical replicate present wi…

cydar mass cytometry

updated 6.5 years ago • kenneth.baker

6 libraries each). In these treatments I have 2 groups of 3 libraries each which are not exactly replicates but should behave similarly. I want to test mainly for differences between the groups in each treatment. Thanks

edgeR edgeR

updated 11.9 years ago • Ruprecht Kathrin

been entertaining the notion that high Ct values, which are generally used as an indicator of no replication due to various reasons, might actually be a response worth prediction via a binomial regression. So for example

Regression HTqPCR Regression HTqPCR

updated 14.2 years ago • Christopher Howerton

nbsp;                        design=~ Replicate + Condition) dds$Condition <- relevel(dds$Condition, "Ctrl") dds <- DESeq (dds,test = "Wald")   The problem is that I'm getting

deseq2 averagePriorsOverLevels

updated 9.5 years ago • laianavarromartin

<div class="preformatted"> I am working on my metagenomic data sets. I have annotated my metagenome against COG database. I would like to use DESeq to look for the overabundant genes in my site. Here is the problem, I only have one site (one metagenome). I would like to compare this one to different sites (each of these site has no replication too) the count data set looks like this: f…

DESeq DESeq

updated 11.4 years ago • Pet Chiang

preformatted">Dear all, I have a data set consisted of 4 cell types with each cell type having 3 replicates. I find the unique signature genes of each cell types using decideTests (see following). I only want the top, say 20

updated 14.6 years ago • Wendy Qiao

Tis1 T1 V1 Ty2 Tis1 T1 V2 ... For each Type Tissue and Time point i have 3 replicates. I need to create a model like Y ~ Type x Tissue x Time and calculate the contrasts (all possible combinations). I need

limma limma

updated 14.5 years ago • Dimitris Kampas

div class="preformatted"> Dear Gordon, i performed 8 technical replicates with two color oligo array. Particularly i have 3 samples: MUT1, MUT2 and Reference (pool of wild type mice), then my

limma limmaGUI oligo limma limmaGUI oligo

updated 19.7 years ago • bzzandrew@interfree.it

a DEG analysis of a microarray data. I have 9 placebo and 9 treated mouse samples. The 3 replicates for each condition factors: 1. TimeOfTreatment <- "2days", " 7days", "12days" 2. TreatmentType <- "placebo" , "drug" 3. BatchID

limma factorial design matrix microarray

updated 6.4 years ago • senthil.duraikannukailasam

to detect allele-specific expression? I have a count table in which each F1 hybrid (3 biological replicates) has separate counts for the paternal and maternal allele. I am considering implementing a paired design in edgeR

edgeR deseq2 ase allele

updated 10.2 years ago • trianglescout

I am analysing a dataset with two timepoints, two conditions within every timepoint and three replicates for every time-treat combination. Furthermore suppose that the hypotheses of interest are (a) DE within timepoint1

deseq2

updated 9.3 years ago • Koen Van den Berge

I use the limma bioconductor for analysing microarray results. There is on my microarrays three replicates for each spot (thus I use correlation or average to deal with these values). My problem is that we have several probes

Microarray limma Microarray limma

updated 18.6 years ago • Christophe Boutte

Hello, This question is in regards to using DiffBind (or DESeq2) with multiple samples. Currently, I am using DiffBind (version 2.8.0) to find differential peaks from ATAC-seq data. I have 7 different cell types with 1-3 replicates each (15 samples total). I'm looking to identify differential peaks among all 7 cell types in order to find peaks that...version 2.8.0) to find differential peaks fr…

diffbind atac-seq differential analysis deseq2

updated 7.2 years ago • duvaller

<div class="preformatted">Dear all, Recently, I have been working on variance stabilization techniques for cDNA microarrays. I have been studying the papers: I. Rocke and Durbin, (2001), `A model for measurement error for gene expression arrays', J. Comput. Biol, 8, 557--569 and II. Durbin, Hardin, Hawkins, and Rocke, (2002), `A variance- stabilizing transformation for gene-expression mi…

Microarray Microarray

updated 20.7 years ago • E Motakis, Mathematics

the solution post online is not available for DEXSeq. I am sure the paired of samples are biological replicates. Could you please let me know whether this can be solved in DEXSeq and report the p-values for the exons?   I used

dexseq

updated 8.6 years ago • zhan0949

not treat. For each of the 4 conditions (untreated, A only, B only, A+B), I have three biological replicates, for a total of 12 samples. I would like to identify genes that are synergistically affected in the combination

deseq2

updated 6.4 years ago • mattobu83

div class="preformatted">Hi, I am trying to replicate the data on biogps.com of the Z-score. They explain the source of how they calculated it. However, I am not familiar

updated 13.9 years ago • Murtuza Zair

I'm running RAIN analysis in R and I'm having trouble with the output. I tried contacting the original authors of the package rain but their emails are inaccessible behind a securty wall. I don't understand what some of the options mean, such as "period". I want to see the rhythmicity over 24h periods but I have data for two days. They are not replicates but rather two 24h hour periods with se…

rain

updated 17 months ago • cereyredondo

<div class="preformatted">Hi again, I am trying DiffBind and loaded my data that looks like this: H3K4m3 4 Samples, 13203 sites in matrix (13792 total): ID Tissue Factor Condition Peak.caller Replicate Intervals 1 wt1 Hela H3K4me3 control1 raw 1 14111 2 wt2 Hela H3K4me3 control2 raw 2 13771 3 treat1 Hela H3K4me3 condition1...4 S…

Genetics DiffBind Genetics DiffBind

updated 13.3 years ago • António Miguel de Jesus Domingues

to look for DE genes in a dataset of 15 samples with 3 conditions (C, A, and B). C: control (5 replicates) A: drug1 treatment (5 replicates) B: drug2 treatment (5 replicates) I found few or no gene with padj<0.1 for C vs. A, and

deseq2

updated 9.4 years ago • qys2007

div class="preformatted">Dear Juan Lin, Only one of your samples has a technical replicate, and this is not enough information to estimate the duplicate (within sample) correlation, so you can't use the duplicateCorrelation...function. You need to either average the two technical replicates (there is a function avearrays in the developmental version of limma for this purpose), or just choose…

affy limma affy limma

updated 15.3 years ago • Gordon Smyth

I have the GNF dataset with Affy expression data from 61 mouse tissues (each with 1 biological replicate, 122 total CEL files) In the end I would like to obtain, for each tissue, the gene list sorted according to the specificity...are, other than trying to get general suggestions: 1) at what point do I use the biological replicates? 2) is there a package that I can use to obtain "relative" exp…

probe affy gcrma probe affy gcrma

updated 18.7 years ago • Cei Abreu-Goodger

were infected or not (inf) and then 3 time points were take and sequenced. Most samples have 2 biol replicates and some 4 biol replicates. My question is the following: I would like to make a contrast were I compare the interaction

updated 11.5 years ago • Maximo Rivarola

div class="preformatted">Hello, I had arrays with 4 replicate spots per gene. I used limma package for data analysis. > targets SlideNumber FileName Cy3 Cy5 Name Field1 1 13617731...WBM WC Field5 After read in data, normalization, I used the following codes for within-array replicate spots. design <- modelMatrix(targets, ref="WC") corfit <- duplica…

Normalization limma Normalization limma

updated 18.2 years ago • Tiandao Li

normalize by or across all treatments I somewhat disagree--If you have meaningful contrasts (with 3 replicates) they will survive normalising all together. If you normalise separately you will almost certainly introduce...25 AM To: bioconductor@stat.math.ethz.ch Subject: [BioC] normalize by or across all treatments In replicates of 3, I have a set of control and 4 treatment arrays. My question…

Normalization Normalization

updated 22.6 years ago • Stephen Henderson

Hi Bioconductor ! I have a time course RNAseq data from 8 time-points with 3 replicates per sample. These data were aligned with STAR, I get the featureCount output. With these file, I wan't to use MasigPro...post to understand that the RNAseq libraries follow this model due to differences between biological replicates inducing that variance differs from mean. I've got a table with samples in c…

R masigpro edgr rnaseq negative binomial model

updated 7.9 years ago • nicolas.hipp

div class="preformatted">Hello, I had arrays with 4 replicate spots per gene. I used limma package for data analysis. > targets SlideNumber FileName Cy3 Cy5 Name Field1 1 13617731...WBM WC Field5 After read in data, normalization, I used the following codes for within-array replicate spots. design <- modelMatrix(targets, ref="WC") corfit <- duplic…

Normalization limma Normalization limma

updated 18.2 years ago • Tiandao Li

is analagous to having duplicate probes for microarrays). So what I want to do is split SGAs by each replicate, giving four times as many samples in each batch and making the data a lot more suitable for batch effect normalisation...etc.) why this would be inappropriate to do? I can't think of any reason why not, since all 20 replicates are part of the same batch, and so should all share the same…

combat sva sga combat batch effect

updated 9.7 years ago • John Townsend

The total genes are 22,000.  Anyway, the point here is that the dataset has only one biological replication from each Cell-line as you can see and only one biological replication for each treatment. Question \#1 : Currently

pathways statistics microarray

updated 8.0 years ago • arronar

series with two conditions (SD and LD) and 3 time points (0, 2, and 4). There are three biological replicates per sample. I have used DESeq2 to do a likelihood ratio test on the following designs:  <pre> full ~ condition + time...between time points 2 and 4? This is my sample table: <pre> sample condition time replicate LD_ZT4_1 LD_ZT4_1 LD 4 …

rna-seq deseq2

updated 4.8 years ago • gtho123

I have 2 genotypes (tolerant and susceptible) and 2 conditions (drought and well-watered) with 4 replications each, essentially a 2x2 factorial experiment with 4 replications. One of my main objectives is to identify drought

deseq2 ANOVA design interaction term

updated 7.9 years ago • tarun2

1) I am working with a multi-factor experiment that, in a sense, doesn't have biological replicates (replicates were done at different times, and the differences in the chemistry/instrumentation is enough to be

updated 13.7 years ago • Ingrid Lindquist

the `VST`-transformed data of the top highly expressed genes from bulkRNAseq across biological replicates (donors), and in a separate plot, visualise the `SCT`-transformed expression of DEGs of this cluster (that also present

Seurat RNASeq DESeq2 scRNAseq

updated 11 months ago • Quang NN

Hi, I'm trying to overlay coverage and sashimi plots for group of RNA-Seq samples (replicates) so that I can have multiple treatment visualized in one simple plot with Gviz. I need to set the coverage ylims...Hi, I'm trying to overlay coverage and sashimi plots for group of RNA-Seq samples (replicates) so that I can have multiple treatment visualized in one simple plot with Gviz. …

Coverage Gviz sashimi

updated 15 months ago • Marco

peaks as a bed file at the sample file generation stage under the peaks column for each sample (3 replicates & 2 conditions) Thank you in advance, Anthony

DiffBind

updated 3.3 years ago • doherta6