I'm also in the same boat as Rich. I run a new bioinformatics core
here and I'm building a pipeline for RNA-seq. Cufflinks for some time
has supported biological replicates, and I'm also curious about the
relative merits of using
bowtie/tophat-cufflinks-cuffmerge-cuffdiff-cummeRbund versus using
tophat-HTSeq?-customScriptForCreatingMatrix?-DESeq. Cufflinks also
gives me a host of other tests (differential splicing load,
differential TSS usage, differential coding output, etc), which also
seem useful for certain applications.
On a related note, does anyone have a workflow for taking multiple bam
files, running HTSeq-count (or another program), plus some other
program or custom script, to produce a matrix of counts as input to
DESeq?
Stephen
-----------------------------------------
Stephen D. Turner, Ph.D.
bioinformatics at virginia.edu
Bioinformatics Core Director
University of Virginia School of Medicine
bioinformatics.virginia.edu
On Tue, Feb 14, 2012 at 6:00 AM, <bioconductor-request at="" r-project.org=""> wrote:
>
> Message: 6
> Date: Mon, 13 Feb 2012 09:28:10 -0800
> From: "Tim Triche, Jr." <tim.triche at="" gmail.com="">
> To: Richard Friedman <friedman at="" cancercenter.columbia.edu="">
> Cc: Bioconductor mailing list <bioconductor at="" r-project.org="">
> Subject: Re: [BioC] DeSeq vs current version of Cuffdiff
> Message-ID:
> ? ? ? ?<cac+n9bwr30evfz5rh7hpnf-k9uq=b4n_sqgr0b49xgmw89jbmg at="" mail.gmail.com="">
> Content-Type: text/plain
>
> Not directly relevant to gene-level RNA-seq DE calls, but rather for
> exon-level DE,
> I found it useful to read this:
>
http://precedings.nature.com/documents/6837/version/1
> In particular, section 4.3 on page 11, and supplementary figures S7
and S8
> on page 19.
>
> I was informed by a coworker that since everyone uses
> BowTie-TopHat-Cufflinks-Cuffdiff, it is the sensible thing to do.
> Conversations with people who know what they are doing (Terry Speed
&
> BCGSC) suggest the matter is not yet settled.
> So I retrieved ~1TB of BAMs, extracted the reads, and started
looking into
> how that compares to DEXSeq and/or subread.
>
> It would be incredibly informative if the Cufflinks and DEXSeq
authors had
> time to weigh in on their strengths/weaknesses. DEXSeq & cummeRbund
both
> offer nice tools for exploring the results; I am curious which
pipeline
> fits best for my needs.
>
> Thanks for bringing this up.
>
>
> On Mon, Feb 13, 2012 at 8:01 AM, Richard Friedman <
> friedman at cancercenter.columbia.edu> wrote:
>
> > Dear Bioconductor list,
> >
> > ? ? ? ?Sometime ago Simon Anders explained the difference
> > between DeSeq and Cuffdiff as follows:
> >
> > "If you have two samples, cuffdiff tests, for each transcript,
whether
> > there is evidence that the concentration of this transcript 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 whether the change is significant
compared to
> > changes in each condition.
> >
> >
http://cufflinks.cbcb.umd.edu/**howitworks.html#hdif<http: cuffli="" nks.cbcb.umd.edu="" howitworks.html#hdif="">
> >
> >
http://cufflinks.cbcb.umd.edu/**howitworks.html#reps<http: cuffli="" nks.cbcb.umd.edu="" howitworks.html#reps="">
> >
> > Can someone please comment on the relative merits of Cuffdiff and
> > DeSeq. I ask here because our sequencing core delivers results
> > based on Cuffdiff and I want to know if I should redo it using
> > DeSeq,I would greatly appreciate any guidance in this matter.
> >
> > Thanks and best wishes,
> > Rich
> > ------------------------------**------------------------------
> > Richard A. Friedman, PhD
> > Associate Research Scientist,
> > Biomedical Informatics Shared Resource
> > Herbert Irving Comprehensive Cancer Center (HICCC)
> > Lecturer,
> > Department of Biomedical Informatics (DBMI)
> > Educational Coordinator,
> > Center for Computational Biology and Bioinformatics (C2B2)/
> > National Center for Multiscale Analysis of Genomic Networks
(MAGNet)
> > Room 824
> > Irving Cancer Research Center
> > Columbia University
> > 1130 St. Nicholas Ave
> > New York, NY 10032
> > (212)851-4765 (voice)
> > friedman at cancercenter.**columbia.edu <friedman at="" cancercenter.columbia.edu="">
> > http://cancercenter.columbia.**edu/~friedman/<http: cancercenter.="" columbia.edu="" ~friedman=""/>
> >
> > I am a Bayesian. When I see a multiple-choice question on a test
and I
> > don't
> > know the answer I say "eeney-meaney-miney-moe".
> >
> > Rose Friedman, Age 14
> >
