[BioC] RNA degradation plot with oligo package GeneFeatureSet objects
heyi xiao
xiaoheyiyh at yahoo.com
Sat Aug 17 00:28:00 CEST 2013
Hi Christian,
Great, I will check that out. Thanks for the info.
Heyi
--------------------------------------------
On Fri, 8/16/13, cstrato <cstrato at aon.at> wrote:
Subject: Re: [BioC] RNA degradation plot with oligo package GeneFeatureSet objects
Cc: bioconductor at r-project.org
Date: Friday, August 16, 2013, 4:32 PM
Dear Heyi,
Yes, xps should work with the Ovine Gene 1.1 ST arrays,
since it does
work with the Human, Mouse and Rat Gene 1.1 ST arrays. These
arrays use
the PGF-files from Affymetrix and
'xps/examples/script4schemes.R' shows
you how to create the 'scheme' files for xps.
Best regards,
Christian
On 8/16/13 9:36 PM, heyi xiao wrote:
> Hi Christian,
> Thanks for the suggestion and sharing of degradation
plot experience. Does xps work with the Ovine Gene 1.1 ST
array? I don’t have a CDF package.
> Heyi
>
> --------------------------------------------
> On Fri, 8/16/13, cstrato <cstrato at aon.at>
wrote:
>
> Subject: Re: [BioC] RNA degradation
plot with oligo package GeneFeatureSet objects
> To: "James W. MacDonald" <jmacdon at uw.edu>,
> Cc: bioconductor at r-project.org
> Date: Friday, August 16, 2013, 2:01
PM
>
> Dear Heyi,
>
> Function 'plotAffyRNAdeg()' of package
'xps' does allow you
> to plot RNA
> degradation plots for Whole Genome and
Exon arrays, see
> Cahpter 5.4.1 of
> vignette 'xps.pdf'.
>
> I have done this for a couple of
HuGene array data, and the
> results look
> interestingly. Especially there is a
difference when you
> compare RNA
> degradation plots from frozen tissues
vs paraffin embedded
> tissues.
> However, I am not sure how to
interpret the results.
>
> Best regards,
> Christian
> _._._._._._._._._._._._._._._._._._
> C.h.r.i.s.t.i.a.n S.t.r.a.t.o.w.a
> V.i.e.n.n.a
> A.u.s.t.r.i.a
> e.m.a.i.l:
cstrato at aon.at
> _._._._._._._._._._._._._._._._._._
>
>
>
> On 8/16/13 6:52 PM, James W. MacDonald
wrote:
> > Hi Heyi,
> >
> > On 8/14/2013 4:47 PM, heyi xiao
wrote:
> >> Hi all,
> >> In affy package, I can use
AffyRNAdeg and
> plotAffyRNAdeg to plot and
> >> check RNA degradation. Is
there any way to do so in
> oligo package for
> >> GeneFeatureSet,which is
equivalent to AffyBatch in
> affy package. I
> >> look at the GeneFeatureSet
and AffyBatch, they
> quite similar. But not
> >> sure what can be done here. I
can either modify
> AffyRNAdeg and
> >> plotAffyRNAdeg functions to
fit them for
> GeneFeatureSet, or I can
> >> convert GeneFeatureSet to
AffyBatch and use the
> affy package
> >> degradation functions. Any
suggestions would be
> highly appreciated.
> >
> > While I suppose you could
hypothetically do the
> conversion, I wonder if
> > it makes conceptual sense.
> >
> > The 3'-biased Affy arrays were
all based off an
> oligo-dT primer that was
> > used to convert mRNA to cDNA, so
the reverse
> transcription proceeded
> > from the 3' end of the mRNA,
always. In this case you
> can wonder about
> > two things. First, how far did
the RT step proceed? Did
> you in general
> > get good RT all the way to the
most 5' of the probes in
> the probesets?
> >
> > Second, since we were using the
polyA tail at the 3'
> end, by definition
> > the mRNA wasn't degraded from the
3' end. However, it
> might have had
> > more or less extensive
degradation from the 5' end, so
> the RT may have
> > gone to completion, but the
degradation had proceeded
> past the most 5'
> > probes.
> >
> > So both things are confounded, as
we cannot distinguish
> RT that didn't
> > proceed too far from highly
degraded mRNA, but no
> matter. What we could
> > do is say how much signal we were
getting from the more
> 5' probes, and
> > decide if we wanted to do
something about that (like
> only use the first
> > 8 probes or whatever).
> >
> > For the newer generation of Affy
arrays, we use a
> random primer, so the
> > RT step proceeds from a random
point in the transcript
> and proceeds
> > towards the 5' end (at least I
think it is still
> directional). Since the
> > RT no longer starts from one end
of the transcript, it
> is no longer
> > clear what differential amounts
of probe signal would
> actually signify.
> >
> > In addition, with the newer
generation of Affy arrays,
> we can collapse
> > the probes into different
probesets, depending on what
> we are trying to
> > measure (e.g., you can try to
measure expression at the
> exon level or
> > the transcript level).
> >
> > I think trying to do this would
be more difficult than
> it would be
> > worth, especially given that I
don't know what you
> would do if you were
> > to decide there had been
degradation.
> >
> > Best,
> >
> > Jim
> >
> >
> >> Heyi
> >>
> >>
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>
>
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