SEQanswers

Go Back   SEQanswers > Bioinformatics > Bioinformatics



Similar Threads
Thread Thread Starter Forum Replies Last Post
DESeq2 Simon Anders Bioinformatics 123 07-06-2015 02:45 AM
DESeq V DESeq2 sebastion RNA Sequencing 35 10-16-2014 07:04 AM
DESeq2 SummarizedExperiment help sindrle Bioinformatics 10 10-23-2013 05:20 AM
Binary characters in cuffcompare result & Questions on cuffdiff nkwuji RNA Sequencing 3 11-03-2010 03:01 AM
Strange cuffdiff result (disagreement between tracking and diff files) jebe Bioinformatics 0 07-01-2010 11:42 AM

Reply
 
Thread Tools
Old 10-19-2013, 06:55 PM   #1
sindrle
Senior Member
 
Location: Norway

Join Date: Aug 2013
Posts: 266
Default No result with DESeq2, but with Cuffdiff 2.1.1

I have a question, I get no significantly changed genes running DESeq2. I have two groups at two time points (paired) and ran DEseq2 v1.2. DESeq2 test the difference between groups, disregarding time point, difference between time points disregarding groups & finally if one group had another effect between the two timepoints than the other.

I have also used Cuffdiff 2.1.1 testing difference between groups, disregarding time point & difference between time points disregarding groups (Cuffdiff cannot handle the last test).

Cuffdiff found over 1000 diff exp. genes and running GAGE it all made biological sense. So why cant I find anything with DEseq2?
sindrle is offline   Reply With Quote
Old 10-20-2013, 09:25 AM   #2
sindrle
Senior Member
 
Location: Norway

Join Date: Aug 2013
Posts: 266
Default

I haw tweeked abit and the results now looks ok, but differing still from cuffdiff. My study design is however one that cuffdiff cannot handle..
sindrle is offline   Reply With Quote
Old 10-21-2013, 01:21 AM   #3
Simon Anders
Senior Member
 
Location: Heidelberg, Germany

Join Date: Feb 2010
Posts: 994
Default

If you told us what you actually did, we might be able to tell you whether it is correct. So, how have you called DESeq2?
Simon Anders is offline   Reply With Quote
Old 10-21-2013, 01:25 AM   #4
sindrle
Senior Member
 
Location: Norway

Join Date: Aug 2013
Posts: 266
Default

Hi!
With this run I get some genes sig. changed and I guess they are more to be trusted than Cuffdiff results due to correct design?
I will see if EdgeR also gives the same results.

library(DESeq2);
library(GenomicFeatures);
library(htSeqTools);
sampleFiles <- list.files(path="/Volumes/timemachine/HTseq_DEseq2",pattern="*.txt");
sort(sampleFiles,decreasing=TRUE);
status <- factor(c(rep("Diabetic",22), rep("Healthy",26)), levels=c("Diabetic","Healthy"));
timepoints = as.factor(c(1,1,1,1,1,1,1,1,1,1,1,2,2,2,2,2,2,2,2,2,2,2,1,1,1,1,1,1,1,1,1,1,1,1,1,2,2,2,2,2,2,2,2,2,2,2,2,2));
patients <- factor(c(88,87,86,83,82,81,75,72,71,70,13,08,01,88,87,86,83,82,81,75,72,71,70,13,08,01,79,77,76,73,67,62,61,55,53,21,04,79,77,76,73,67,62,61,55,53,21,04));
sampleTable <- data.frame(sampleName = sampleFiles, fileName = sampleFiles, status=status, timepoints=timepoints, patients=patients);
directory <- c("/Volumes/timemachine/HTseq_DEseq2/");
des <- formula(~patients + timepoints*status);
ddsHTSeq <- DESeqDataSetFromHTSeqCount(sampleTable = sampleTable, directory = directory, design= des);
ddsHTSeq;
dds <- estimateSizeFactors(ddsHTSeq);
dds <- estimateDispersions(dds);
dds <- nbinomWaldTest(dds, maxit = 5000)
resultsNames(dds);
statusResults <- results(dds, "status_Healthy_vs_Diabetic");
timepointsResults <- results(dds, "timepoints_2_vs_1");
statusTreatmentResults <- results(dds, "timepoints2.statusHealthy");
write.csv(as.data.frame(statusResults),file="/Volumes/timemachine/HTseq_DEseq2/1_Results/statusResults.csv");
write.csv(as.data.frame(timepointsResults),file="/Volumes/timemachine/HTseq_DEseq2/1_Results/timepointsResults.csv");
write.csv(as.data.frame(statusTreatmentResults),file="/Volumes/timemachine/HTseq_DEseq2/1_Results/statusTreatmentResults.csv");
plotDispEsts(dds)

Last edited by sindrle; 10-21-2013 at 01:30 AM.
sindrle is offline   Reply With Quote
Reply

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 02:54 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2020, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO