Thats fantastic. It feels good when I understand whats happening under the
hood.

Thanks so much Mark.

Gowthaman

On Fri, Jun 22, 2012 at 2:50 AM, Mark Robinson <mark.robinson@imls.uzh.ch>wrote:

> Hi Gowthaman,
>
> You shouldn't manually specify the offset in glmFit(), unless you have a
> specific need to.  Short answer, you should use:
>
> fit <- glmFit(d, design)
>
>
> >>>> Lib Fe+.1 has only 4 million reads while other are 9 million +. But
> >>>> still the norm.factors are not much different. With my naive
> >>>> understanding i expect Fe+.1 to be very different from others. I would
> >>>> like to know if what I see is okay?
>
> This is ok, since the offset used in the downstream modeling is actually
> the product of the lib.size and norm.factors columns.
>
> Best,
> Mark
>
> ----------
> Prof. Dr. Mark Robinson
> Bioinformatics
> Institute of Molecular Life Sciences
> University of Zurich
> Winterthurerstrasse 190
> 8057 Zurich
> Switzerland
>
> v: +41 44 635 4848
> f: +41 44 635 6898
> e: mark.robinson@imls.uzh.ch
> o: Y11-J-16
> w: http://tiny.cc/mrobin
>
> ----------
> http://www.fgcz.ch/Bioconductor2012
>
> On 22.06.2012, at 11:31, gowtham wrote:
>
> > Hi Belinda,
> > I think, i am bit confused now. The help document suggest, i should use
> > only one of "offset" and "lib.size". Seems like both of them take the
> > library size into account. And sounds like "offset" has a preference when
> > both are supplied.
> >
> > So, my question is do I have to explicitly ask for one or other? And do I
> > have to explicitly give it a value?
> >
> >
> > fit <- glmFit(d, design)
> >
> > OR
> >
> >
> > fit <- glmFit(d, design, offset=NULL)
> >
> > OR
> >
> > fit <- glmFit(d, design, lib.size=c(9664343, 11248827, 4194124, 9963626))
> >
> > should I supply some values for "lib.sizes". Note, my DGEList already has
> > library size information in it.
> >
> >
> > Once again thanks for your answer and pointer to glmFit.
> > Gowthaman
> >
> > On Fri, Jun 22, 2012 at 2:18 AM, gowtham <ragowthaman@gmail.com> wrote:
> >
> >> Thanks very much Belinda. That is comforting.
> >>
> >> My DGEList object has library sizes added to it. Do I still need to
> supply
> >> a numeric vector with library sizes while fiting glm? Or is it
> >> automatically pulled from DGEList object?
> >>
> >> Reading help, i understand its automatic. Please advice me if I am
> wrong.
> >> " If y is a DGEList object then the default for lib.size is the product
> >> of the library sizes and the normalization factors (in the samples slot
> >> of the object). "
> >>
> >> Thanks,
> >> Gowthaman
> >>
> >>
> >>
> >>
> >> On Thu, Jun 21, 2012 at 4:58 PM, Belinda Phipson <phipson@wehi.edu.au
> >wrote:
> >>
> >>> Hi Gowthaman
> >>>
> >>> Your output looks fine. What is more important is that library size is
> >>> taken into account as an offset later on when you fit the glm. See
> >>> help(glmFit).
> >>>
> >>> Cheers,
> >>> Belinda
> >>>
> >>> -----Original Message-----
> >>> From: bioconductor-bounces@r-project.org [mailto:
> >>> bioconductor-bounces@r-project.org] On Behalf Of gowtham
> >>> Sent: Friday, 22 June 2012 9:40 AM
> >>> To: bioconductor
> >>> Subject: Re: [BioC] edgeR: calcNormFactors question
> >>>
> >>> Sorry about repeated mailing: I have attached a smear plot of the data
> >>> incase that helps anyone attempting to answer my doubt.....
> >>>
> >>>
> >>> On Thu, Jun 21, 2012 at 4:07 PM, gowtham <ragowthaman@gmail.com>
> wrote:
> >>>
> >>>> Hi Everyone,
> >>>> I am analyzing a RNAseq experiment with two groups each having two
> >>>> replicates. One out of 4 libraries have only half as much reads
> >>>> mapping to genome.
> >>>>
> >>>> Lib Fe+.1 has only 4 million reads while other are 9 million +. But
> >>>> still the norm.factors are not much different. With my naive
> >>>> understanding i expect Fe+.1 to be very different from others. I would
> >>>> like to know if what I see is okay?
> >>>>
> >>>>> oldsetDGE <- calcNormFactors(oldsetDGE) oldsetDGE$samples
> >>>>      group lib.size norm.factors
> >>>> fe-.1     2  9664343    0.9865411
> >>>> fe-.2     2 11248827    1.0812947
> >>>> fe+.1     1  4194124    0.9662389
> >>>> fe+.2     1  9963626    0.9701888
> >>>>
> >>>>
> >>>> Thanks very much,
> >>>> Gowthaman
> >>>> --
> >>>> Gowthaman
> >>>>
> >>>> Bioinformatics Systems Programmer.
> >>>> SBRI, 307 West lake Ave N Suite 500
> >>>> Seattle, WA. 98109-5219
> >>>> Phone : LAB 206-256-7188 (direct).
> >>>>
> >>>
> >>>
> >>>
> >>> --
> >>> Gowthaman
> >>>
> >>> Bioinformatics Systems Programmer.
> >>> SBRI, 307 West lake Ave N Suite 500
> >>> Seattle, WA. 98109-5219
> >>> Phone : LAB 206-256-7188 (direct).
> >>>
> >>>
> >>> ______________________________________________________________________
> >>> The information in this email is confidential and intended solely for
> the
> >>> addressee.
> >>> You must not disclose, forward, print or use it without the permission
> of
> >>> the sender.
> >>> ______________________________________________________________________
> >>>
> >>
> >>
> >>
> >> --
> >> Gowthaman
> >>
> >> Bioinformatics Systems Programmer.
> >> SBRI, 307 West lake Ave N Suite 500
> >> Seattle, WA. 98109-5219
> >> Phone : LAB 206-256-7188 (direct).
> >>
> >
> >
> >
> > --
> > Gowthaman
> >
> > Bioinformatics Systems Programmer.
> > SBRI, 307 West lake Ave N Suite 500
> > Seattle, WA. 98109-5219
> > Phone : LAB 206-256-7188 (direct).
> >
> >       [[alternative HTML version deleted]]
> >
> > _______________________________________________
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> > Search the archives:
> http://news.gmane.org/gmane.science.biology.informatics.conductor
>
>


-- 
Gowthaman

Bioinformatics Systems Programmer.
SBRI, 307 West lake Ave N Suite 500
Seattle, WA. 98109-5219
Phone : LAB 206-256-7188 (direct).

	[[alternative HTML version deleted]]

