[BioC] Limma and time-course data

Gordon Smyth smyth at wehi.edu.au
Sat Mar 4 11:34:13 CET 2006 

Dear Mick,

The JSM workshop example was designed to illustrate a particular data 
analysis for which choosing a set number of genes was appropriate.

I am not clear why you're restricting your reading to the workshop 
example. Is there any reason why you're not also looking at Section 
8.8 of the User's Guide titled "Time Course Experiments"? Although 
that section is not very long, it does answer several of the 
questions that you've been asking. It shows how to construct 
different F-statistics for different purposes, how to extract the 
p-values, and how to select genes for a given p-value cut-off.

BTW, I note that Naomi told you a couple of days ago how to get the p-values:

   https://www.stat.math.ethz.ch/pipermail/bioconductor/2006-March/012130.html

Was Naomi's answer too brief for you to be able to use?

Best wishes
Gordon

At 02:23 AM 4/03/2006, michael watson \(IAH-C\) wrote:
>Hi Guys
>
>OK, so I have access to the F statistic, but the example at
>http://bioinf.wehi.edu.au/marray/jsm2005/lab5/lab5.html simply uses the
>F statistic to find the top 500 changing genes - somewhat arbitrary.
>
>Can I access p-values for these F statistics, so that I can look at
>genes where time is a significant factor at, say, p<=0.05?
>
>Many thanks
>Mick
>
>-----Original Message-----
>From: Naomi Altman [mailto:naomi at stat.psu.edu]
>Sent: 02 March 2006 19:21
>To: michael watson (IAH-C); Gordon Smyth
>Cc: bioconductor at stat.math.ethz.ch
>Subject: Re: [BioC] Limma and time-course data
>
>Usually you would fit a factorial design and compile the contrasts into
>3 F-tests: time, strain and interaction.
>
>--Naomi
>
>At 12:25 PM 3/2/2006, michael watson (IAH-C) wrote:
> >Thanks for your help, but one more question: if I have two factors, one
>
> >of which is time, and the other is strain, and say I have used a common
>
> >reference, there should be two F tests, one for time and one for
> >strain, no?
> >
> >________________________________
> >
> >From: Gordon Smyth [mailto:smyth at wehi.edu.au]
> >Sent: Thu 02/03/2006 1:45 AM
> >To: michael watson (IAH-C)
> >Cc: bioconductor at stat.math.ethz.ch
> >Subject: RE: [BioC] Limma and time-course data
> >
> >
> >
> >You will need at least as many contrasts as time points minus one. So
> >if you have 3 times, you need at least 2 contrasts. It is just like
> >oneway anova.
> >
> >Cheers
> >Gordon
> >
> >At 09:39 AM 2/03/2006, michael watson \(IAH-C\) wrote:
> > >Hi Gordon
> > >
> > >And a lightbulb goes on just above my head!  It was beginning to
> > >confuse me where the F test came in.
> > >
> > >So once I have fit the contrasts (given that they span the entire
> > >time course, so if I have endpoint-startpoint as default, I will be
> > >OK) I can access the F statistic through the fitted model object?
> > >And this has a significance value associated with it?
> > >
> > >Thanks alot :)
> > >
> > >Mick
> > >
> > >________________________________
> > >
> > >From: Gordon K Smyth [mailto:smyth at wehi.EDU.AU]
> > >Sent: Wed 01/03/2006 9:59 PM
> > >To: michael watson (IAH-C)
> > >Cc: bioconductor at stat.math.ethz.ch
> > >Subject: [BioC] Limma and time-course data
> > >
> > >
> > >
> > >Dear Mike,
> > >
> > >limma does exactly what you want.  The approach outlined in the
> > >User's Guide (and in the workshop)
> > >  finds genes which change over time without worrying about which
> > >particular time the genes differ at.  I think you may have missed the
>
> > >fact that the approach uses the F-test, not the individual contrast
> > >p-values.  You will get the same F-test regardless of how you specify
>
> > >the contrasts, as long as the contrasts span all the times.
> > >
> > >As far as verbosity is concerned, limma is a general purpose program,
>
> > >not specifically for time courses.  So to create the F-test, you do
> > >need to explicitly setup a set of contrasts.  Strictly speaking, I
> > >could get limma to make a set of contrasts automatically if it is
> > >known that you want to do an F-test.  But making the contrasts takes
> > >only a few lines of code (as you show below), so I can live with
> > >that, at least for now.
> > >
> > >Cheers
> > >Gordon
> > >
> > > > Date: Tue, 28 Feb 2006 12:30:47 -0000
> > > > From: "michael watson \(IAH-C\)" <michael.watson at bbsrc.ac.uk>
> > > > Subject: [BioC] Limma and time-course data
> > > > To: <bioconductor at stat.math.ethz.ch>
> > > >
> > > > Hi
> > > >
> > > > Googling the list shows this up to be a rather hot topic, but I
> > > > just wanted to ask a few more questions.
> > > >
> > > > Firstly, it seems the plan for tackling time course data within
> > > > limma is to treat each time-point/treatment combination as a
> > > > coefficient to be estimated.  Thus, to ask "which genes are
> > > > changing over time", one must fit contrasts that compare every
> > > > single time point to every other time point, pairwise, and look
> > > > for any gene that is significant in one or more of those
>comparisons.  Is that correct?
> > > >
> > > > I am also a tad confused by the documentation, which states (on
> > > > page
> > > > 47):
> > > >
> > > > "> cont.wt <- makeContrasts(
> > > > + "wt.6hr-wt.0hr",
> > > > + "wt.24hr-wt.6hr",
> > > > + levels=design)
> > > >> fit2 <- contrasts.fit(fit, cont.wt)
> > > >> fit2 <- eBayes(fit2)
> > > >
> > > > Any two contrasts between the three times would give the same
>result.
> > > > The same gene list
> > > > would be obtained had "wt.24hr-wt.0hr" been used in place of
> > > > "wt.24hr-wt.6hr" for example."
> > > >
> > > > I'm confused why "wt.24hr-wt.0hr" and "wt.24hr-wt.6hr" would give
> > > > the same gene list.  The first looks for differences in wt between
>
> > > > time points 0 and 24, and the second looks for differences between
>
> > > > time points 6 and 24.
> > > >
> > > > I guess, to me, this all seems a bit verbose and difficult,
> > > > particularly for large time-course experiments where many
> > > > biologists want to subset their data to those genes that change
> > > > over time and thus want to ask the question "does time have an
> > > > effect on the expression of my gene?" and are not particularly
> > > > bothered, at this stage, which particular time points those genes
>differ at.
> > > >
> > > > Thanks in advance
> > > >
> > > > Mick
> > >
> > >
> > >
> > >
> >
> >_______________________________________________
> >Bioconductor mailing list
> >Bioconductor at stat.math.ethz.ch
> >https://stat.ethz.ch/mailman/listinfo/bioconductor
>
>Naomi S. Altman                                814-865-3791 (voice)
>Associate Professor
>Dept. of Statistics                              814-863-7114 (fax)
>Penn State University                         814-865-1348 (Statistics)
>University Park, PA 16802-2111

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