Bioconductor Forum

appreciate any guidance on this matter. Take into account that batch correction is needed since the replicates for the same condition where extracted on different days (batches

rnaseqDTU Normalization

updated 6 months ago • Samuel

Hello! I am looking for DE genes comparing condition1 vs control. I have 3 biological replicates per each case, so 6 in total. I am using this design: > design=~ sex + condition and I am filtering low expressed genes

RNASeq DESeq2 RNASeqData

updated 15 months ago • bioshot.com

expression analysis between different treatments.I have 4 conditions and 4 four biological replicates for each conditions. I’ve performed differential expression analysis using DESeq2 building a dds model considering

DESeq2

updated 4.1 years ago • giulia.lopatriello

Hi, I have a dataset of ribo-depleted RNASeq from 3 pairs of cell populations (with replicates). We have reasons to believe there will be abnormal expression at specific genomic regions in some of those populations

DESeq2 RNASeq

updated 3.9 years ago • anatmela

have a time course RNAseq experiment with 2 different genotypes and 2 times (0h and 2h), there are 3 replicates each time I am looking for a way to output genes that are degraded at 2h but not affected by the genotype condition

DESeq2 RNASeqData DifferentialExpression

updated 3.0 years ago • Julie

I am trying to use deseq2 to compare differential gene expression across multiple generations of repeated treatment. I have three replicates for each condition and there is a treated and untreated/control condition for each sample. In total these are my...compare differential gene expression across multiple generations of repeated treatment. I have three replicates for each condition and there is…

rnaseq deseq2

updated 2.8 years ago • lunarskye222

the data beforehand** which is absolutely fine. We are confident that we have enough samples and replicates for any of the subsets. However, the dispersion/size factor estimation takes significantly longer than anticipated

DEXSeq

updated 2.8 years ago • Ben J

<div class="preformatted">Hi, I use limma to analyse my dye swap. I have 3 replicates with a spacing of 1152. When I use the gls.series function I obtain a very small correlation. fit <- gls.series(data...div class="preformatted">Hi, I use limma to analyse my dye swap. I have 3 replicates with a spacing of 1152. When I use the gls.series function I obtain a very small correlatio…

limma limma

updated 21.9 years ago • Valerie Cognat

experiment uses identical RNA except for 42 spike-ins which are varied by condition. There are 3 replicates, making 6 arrays in all. The number of "significant" genes at p<.01 using limma with eBayes (but no multiple comparison

limma limma

updated 19.9 years ago • Naomi Altman

on a 2x2 factorial experiment (2 drought contrasting genotypes and 2 contrasting conditions) with 4 replicates per sample to have a 16 experimental units. Before proceeding to run the differential expression, I initially

deseq2 rlog transformation

updated 8.0 years ago • tarun2

has control (C) and experiment (E) conditions. The screen uses 3 x 96 well plates and C and E have 2 replicates each, hence C and E have 6 plates each in total. It gets complicated because there is a repeat of the whole experiment

cellHTS2 cellHTS2

updated 17.3 years ago • Steve Taylor

as to why it requires the layout ? I didn't think newer array designs had print tip groups or replicate spots ? -------------------------------------- Dario Strbenac Research Assistant Cancer Epigenetics Garvan Institute of Medical Research Darlinghurst

updated 15.0 years ago • Dario Strbenac

<div class="preformatted">I have a short RNAseq time series dataset (two time pointsl T0 and TN) and I have two conditions Undrugged and Drugged. For each of the drug conditions the timepoints are paired (i.e one T0 corresponds to one T1). There are two T0/TN-paired replicates for each drug condition. So we have (Drugged_T0_A, Drugged_T1_A), (Drugged_T0_B, Drugged_T1_B), (Undrugged_T0_C, Un…

RNASeq RNASeq

updated 13.1 years ago • Simon Knott

of all. Are there any pitfalls to do that? In more detail: I have 9 tissue samples (with 3 or 4 replicates) and I'd like to find tissue enhanced genes. So the plan is to perform DE analysis with DESeq2 with a dataset where

deseq2

updated 5.8 years ago • varga.torda

Dear EDGER support team, I have a single factor experiment with the 5 different samples having three replicates each. group <- factor(c("a","a","a","b","b","b","c","c","c","d","d","d","e","e","e")) Using a __GLM based approach,__ and using __TreatDGE__ function, I am trying...EDGER support team, I have a single factor experiment with the 5 different samples having three replicates each…

edger

updated 11.1 years ago • candida.vaz

each of which was studied at 2 time points (T1,T2), under 2 conditions ( drought, water), and with 2 replicates. I performed the analysis like this: 1) Upload counts data for all strains and all conditions (48 columns) to the deseq2

deseq2

updated 7.0 years ago • naktang1

Hi, I am currently performing DE analysis on fungal gene expression in axenic and in planta conditions. I have 3 biological replicates subjected to the two conditions. When inspecting the Dispersion estimates and MA (LFC shrunk) plots, these are the...DE analysis on fungal gene expression in axenic and in planta conditions. I have 3 biological replicates subjected to the two conditions. When ins…

DESeq2

updated 5.1 years ago • hazwanfikri0505

package will do a good job of differential expression between conditions, assuming there are some replicates. Best wishes Gordon > Date: Thu, 12 Feb 2009 10:18:17 +0100 > From: Ingunn Berget <ingunn.berget at="" umb.no=""> > Subject

Genetics Genetics

updated 16.9 years ago • Gordon Smyth

some RNA-seq generated with a plant species. I have a time series experiment with 8 time point and 4 replicates for each time point.   I just have a question concerning the normalisation method used by Deseq2 and his usability

deseq2

updated 7.4 years ago • Alex So

Reading Read Group Info /data/projects/LL/comp/test/04_DE/test_cdout/ read_groups.info Writing replicates Table Reading /data/projects/LL/comp/test/04_DE/test_cdout/genes.fpkm_tracking Error in scan(file, what, nmax

cummeRbund cummeRbund

updated 13.2 years ago • Estefania

<div class="preformatted">Hi, I'm "playing" a bit with the vignette on customMethods in the affy package. If I try to replicate the example (or doing my own) for a new method for summary expression (i.e. "huber"), I get: expresso(mydata,bg.correct=F...Hi, I'm "playing" a bit with the vignette on customMethods in the affy package. If I try to replicate the example (or doing my own) for a …

updated 20.6 years ago • stecalza@tiscali.it

I want to visualize each of my samples (3 different conditions; 20 timepoints - no need to visualize replication because things will become too messy) . But I'm frankly a bit overwhelmed by the plethora of options. My doubts include

updated 17.1 years ago • Yannick Wurm

from MASS package. In help, gls.series is mentioned to be used in hight correlation between spots replication. In my case I dont have this. I visited the help files but I dont get knowledge for decide how I use in my data set. The

limma limma

updated 20.8 years ago • Marcelo Luiz de Laia

melanogaster** to do the analysis; My sample is here, a 6 time points experiment without replicates. I want to compare specific time effect ``` Hours <- c(0,1,2,4,8,16) Time <- paste0(Hours,"hrs") y <- DGEList(counts=x, group = Time

edger

updated 5.6 years ago • Ruixuan

in the meantime the following code should work: d <- DGEList(counts = cD at data, group = cD at replicates) d <- calcNormFactors(d) cD at libsizes <- d$samples$norm.factors Best wishes, Tom Hardcastle -- Dr. Thomas J. Hardcastle

Normalization edgeR baySeq DESeq Normalization edgeR baySeq DESeq

updated 14.2 years ago • Thomas J Hardcastle

or stay flat, etc. To check this, I made a single series (only females) design matrix, with three replicates for each of the three-time points (quadratic regression). Then, I called p.vector() to get a regression fit for each

timecourse rnaseq masigpro

updated 8.1 years ago • jelisaveta.jeki.djordjevic

pseudoalignments.sorted.bam") ribroDat <- readRibodata(ribofiles, rnaFiles = rnafiles, replicates = reps) Here is the error medd\ssage: Reading ribosomal files... ......done! Reading rna files...Error in .Call2("solve_user_SEW0

riboSeqR

updated 6.3 years ago • Hui.Pan

with differential expression at the transcript (isoform) level. I have 6 different conditions (2 replicates/condition). I want to know if I can use DESeq2 for that or if the package can only be used at the gene level. Thanks, Alicia

DESeq2 DESeq2

updated 11.7 years ago • Alicia R. Pérez-Porro

I am analyzing microarray data from three different batches. There are many technical and biological replicates in the batches. In the past, when I had only two batches, I was using removeBatchEffect function from limma, giving

microarray limma removebatcheffect()

updated 9.6 years ago • Eleni Christodoulou

When running DESeq2 with the same expression data I get different results depending on the order of the columns in my count data matrix. Is this expected behaviour or am I messing something up? My code looks like this: <pre> countData <- read.table("matrix") colData <- read.table("samples", header=F) colnames(colData) <- c("", "genotype", "treatment") colData$trea…

deseq2

updated 10.3 years ago • lelle

the given expression data are fully pre-processed including data normalization and averaging of replicates.  As @Michael Love explained in this question <https://support.bioconductor.org/p/87999/> "the normalized

deseq2 rnaseq mfuzz

updated 7.5 years ago • rbenel

that I want to look into their differential gene expression. I call them A, B, C and D. I have 5 - 8 replicates for each group and I am using DESeq2 for the analysis. Which of the following ways is a recommended setting to continue

deseq2 rna-seq r

updated 5.6 years ago • thjnant

treated1 treated2, treated3 vs. control1,control2,control3 50bp data can be seen as technical replicates of 100bp data?cluding two conditions: treated1 treated2, treated3 vs. control1,control2,control3 do u know how

updated 13.3 years ago • wang peter

Ex-1-Test- 02 Ex-1-Test -03 Ex-2-Test-01 Ex-2-Test-02 Ex-2-Test-03 Three samples with biological replicates. I wish to do ANOVA. Other than using limma package, I wish to do ANOVA with some other package especially with multtest

multtest limma multtest limma

updated 17.0 years ago • John Antonydas Gaspar

<div class="preformatted">Dear List, I would like to ask if there is such a bioconductor package available that can help to achieve the following purpose. Thank you very much in advance! I got 16 Affy chips corresponding to 4 samples: wild-type treated, wide-type untreated, knocked-down treated, and knocked-down untreated, i.e. 4 replicates for each sample. I want to look at the expressi…

affy affy

updated 15.5 years ago • Yuan Hao

div class="preformatted">Dear List, I have several biological replicates affy arrayes (a simple one group 4 arrayes), and tried to use eBayes to get the differentially expressed genes. The

affy affy

updated 16.3 years ago • Roger Liu

Dear all, I've a dataset of around 150 RNAseq from different patients and after performing quantification of transcript with Salmon. I'm interested in founding immunologic signature correlate with the expression of a specific gene isoform. So I've retrieve the abundance (TPM) of that isoform among my 150 RNAseq and I'm using this as a continuous phenotype into GSEA. Then I've performed gene su…

deseq2 gsea tpm rnaseq salmon rna seq

updated 7.2 years ago • leo_CD

as_html, options = options) : Start tag expected, '<' not found [4] The code to replicate is below, using bioconductor 3.7 and TCGAbiolinks 2.8.1 project <- 'TCGA-KIRC' query <- list() query$clinical <- GDCquery

tcgabiolinks

updated 7.6 years ago • andre.verissimo

a bit of trouble interpreting a result after running DESeq2 on an RNA-seq dataset with 6 biological replicates for multiple conditions (comparing a parental cell line and multiple independent gene KOs). In particular, for

deseq2

updated 6.8 years ago • algejohnson

i.set], data[i.set]) : need at least four unique 'x' values Does this mean I don't have enough replicates? thanks for the help Andreia -- -------------------------------------------- Andreia J. Amaral Unidade de Imunologia Clínica Instituto de Medicina Molecular

miRNA qPCR miRNA qPCR

updated 15.0 years ago • Andreia Fonseca

Hello edgeR support team, First off, thank you for the incredibly useful and well-documented package. It has been a great help for our research. We have a data set with two tissues sampled from many species, which are clustered into several species groups (orthology assignment is upstream of this analysis). The sampling is unbalanced in both number of biological replicates per species a…

edgeR

updated 4.6 years ago • David

results() with Wald test. I have treated (APO) /untreated (DMSO) sample and 6 timepoint (incl 0) 4 replicate. When I run my script  I've got the same result for every timepoint when I want to compare APO vs DMSO. My aim is to...sampleTable is: <pre> > sampleTable </pre> <pre> sampleName fileName condition replicate timeP 1 APO_0_A APO_0…

timecourse deseq2 multiple factor design LRT

updated 7.1 years ago • juditkovacsbio

updated 20.1 years ago • Ana Conesa

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updated 19.8 years ago • Mike White

Hi, I'm currently exploring methods for batch correction with my RNA-seq dataset from a series of clinical samples. The goal of this approach is to remove batch effects for PCA and other visualization approaches, as well as to generate input for other analytical pipelines that do not include internal methods for batch correction. Our data is comprised of time-courses from individual patients th…

limma batch effect rna-seq

updated 7.3 years ago • Mike A

> In limma manual it is written that correlation should be negative for > dye swaps- why is it positive?- is it a question of wrong model matrix > or is it something wrong with my samples? I concur with Juan Pedro that the most likely cause of this problem is mislabeling of the data; often they are already 'swapped back' for convenience, which is of course wrong. Regarding dye-bias, i…

limma dyebias limma dyebias

updated 15.9 years ago • Philip Lijnzaad

Edit: this was in answer to https://support.bioconductor.org/p/40047/ Dear Florence, Your code looks to be correct. Best wishes Gordon

limma

updated 11.0 years ago • Gordon Smyth

Dear all, referring to scRNA-seq analysis, many thanks to Aaron Lun and Helena Crowell, for writing the following tutorials on OSCA and MUSCAT : https://osca.bioconductor.org/multi-sample-comparisons.html http://biocworkshops2019.bioconductor.org.s3-website-us-east-1.amazonaws.com/page/muscWorkshop__vignette/ 'd appreciate also having your suggestions on the following case of scRNAseq…

scRNAseq

updated 6.0 years ago • Bogdan

This question is because I am misunderstanding how certain things fit together in Limma. There is no example like this in the documentation, and I am trying to figure out how to do this based on examples section 10.5 and 14.1. sorry for the lengthy post, this is a complicated one, but it might be an interesting case example for some of you. A simplified version of my experiment follows Backgr…

limma limma

updated 21.0 years ago • Pita

Dear all! I am trying to analyse a timecourse experiment using two-color microarrays with limma. There are 5 timepoints compared in a loop-design, i.e 1 vs 2, 2 vs 3,?, 4 vs 5, 5 vs 1. This experiment is supposed to be repeated 3 more times (with independent samples) but my collaborators were interested to see the results for this first set of experiments before they hybridise the rest. I have p…

TimeCourse limma limmaGUI timecourse TimeCourse limma limmaGUI timecourse

updated 20.4 years ago • Claus Mayer

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updated 20.0 years ago • Hua Weng

<div class="preformatted">Dear Igoff, I have a trouble to understand some value in cummeRbund - I am sorry for disturbing, but I did not find any answer on Internet. I have two mouses paired-end sequenced. I mapped with Tophat2 - make Cufflinks - and Cufdiff (one replicates - two conditions) - and CummeRbund. It is very nice graphs and outputs, but I would like to get just one number, whi…

cummeRbund cummeRbund

updated 11.7 years ago • Guest User

ref D 16 16 7827_basicSat.gpr D ref Every other slide is a dyeswapped technical replicate and per "group" (A,B,C,D) there are 2 biological replicates. > K=read.maimages(targets$FileName, source="genepix.median

updated 18.2 years ago • dorthe.belgardt@medisin.uio.no

3 3 3 # these are the positions along the axis for the tick marks, technical # replicates from 1 to 3 (replicates of one factor level combination), 4 to 6 # ... > i [1] 3 6 9 12 15 18 21 24 27 30 34 37 39 42 45 49 52 54 57 60 63

affy affy

updated 21.5 years ago • Arne.Muller@aventis.com

and I am also confused on the final results. I have 50,000 genes, 2 conditions, each condition 4 replicates. 25,000 genes have >=8 counts in at least one condition. The total counts in each replicate is as following: cond1_rep1

Normalization edgeR DESeq Normalization edgeR DESeq

updated 14.6 years ago • Xiaohui Wu

200.4 2 20 cellLine_I Melanoma 58.7 255.3 1 I have 2-3 biological replicates (NOT technical replicates) of RNA-seq data for each cell line. What I tried to do with limma is design <- model.matrix

limma duplicateCorrelation

updated 6.7 years ago • Kei Enomoto

but I would lose the genotype structure of the data which would result in a form of pseudo-replication due to including non-independant genotype replicates as independant samples. I would be interested to hear

deseq2 DeSeq2

updated 7.3 years ago • bdy8

Array. The experiment design includes three treatments, namely A, C and G. We have three biological replicates for each treatment, thus A1, A2, A3, C1, C2, C3 and G1, G2, G3. A1, C1 and G1 were from the first batch (microarray experiment...noise between batch) destroyed everything. By accident, I used gcrma to normalize the three replicates from each treatment separately. Something dramatically…

Microarray Normalization Clustering zebrafish probe gcrma Microarray Normalization probe

updated 16.9 years ago • Jun Yin

nbsp; The experimental design is an infection time course with 10 samples (2 biological replicates for each timepoint except 2), 3 timepoints (0h, 12h, 36h) and two strains of mice (WT, and ATG knockout (KO)): 1. WT.0h.a 2. WT.0h.b...I have 3 questions/concerns: 1. Can I perform contrasts with the treatment groups that have no replicates and still get meaningful results? 2. The BCV dist…

edger

updated 10.8 years ago • dsperley

load(con) > dj <- djfrag$dj > fragments <- djfrag$fragments > set.seed(1234) > replicate(2, countOverlaps(dj, fragments)) *** caught segfault *** address 0xa2a7f58, cause 'memory not mapped' Traceback: 1: .Call(.NAME...10: sapply(integer(n), eval.parent(substitute(function(...) expr)), simplify = simplify) 11: replicate(2, countOverlaps…

genomicranges IRanges

updated 9.6 years ago • Stephen Cristiano