[BioC] edgeR: a question about library size

[Mark Robinson]

Hi Naomi.

Well, you don't actually adjust the read counts.  Basically, you are estimating the offset (from all the data) that gets used in the generalized linear model.

Cheers,
Mark

On 2010-06-20, at 1:02 PM, Naomi Altman wrote:

> I do not understand how we can do a multiplicative read count adjustment.
> 
> If K is Poisson(v) then E(K)=Var(K)=v.  If we do twice the sequencing effort, then E(K)=Var(K)=2v.  But if we multiple by 2, then E(2K)=2v and Var(2K)=4v.  So, how can
> this type of adjustment work properly?
> 
> --Naomi
> 
> At 08:15 AM 6/17/2010, Mark Robinson wrote:
>> Hi Raffaele.
>> 
>> In my experience, you're better off with the number of mapped reads.  But, a safer way is to do something data-driven.  For example, TMM normalization (http://genomebiology.com/2010/11/3/R25) is implemented in the calcNormFactors() function.  See also the docs and the user's guide.
>> 
>> Hope that helps.
>> 
>> Cheers,
>> Mark
>> 
>> On 2010-06-17, at 10:00 PM, rcaloger wrote:
>> 
>> > Hi,
>> > I am using edgeR to detect differential expression in NGS experiments.
>> > I have a brief question on what I should considered as "total size of my
>> > libraries".
>> > In my case I have a set of samples that have a quite large variation  in
>> > the library size:
>> >
>> > Total reads Mapped reads
>> >
>> > 1 11076283 8736308
>> >
>> > 2 5881045 4006468
>> >
>> > 3 7139703 5108608
>> >
>> > 4 9089153 5643701
>> >
>> > 5 9723103 8457914
>> >
>> > 6 15570265 8706332
>> >
>> > 7 15844448 12056310
>> >
>> > 8 13375681 8663496
>> >
>> > 9 14997114 8799752
>> >
>> > 10 15744584 8555922
>> >
>> > 11 4642056 3201515
>> >
>> > 12 6458028 4277204
>> >
>> > 13 13206724 9466118
>> >
>> > 14 3035032 2148730
>> >
>> >
>> > Should I insert as lib.size parameter the values referring to the real
>> > size of the libraries (Total reads) or
>> > simply the size of the mapped reads (Mapped reads)
>> >
>> > Thanks for the help
>> > Raffaele
>> >
>> > --
>> >
>> > ----------------------------------------
>> > Prof. Raffaele A. Calogero
>> > Bioinformatics and Genomics Unit
>> > Dipartimento di Scienze Cliniche e Biologiche
>> > c/o Az. Ospedaliera S. Luigi
>> > Regione Gonzole 10, Orbassano
>> > 10043 Torino
>> > tel.   ++39 0116705417
>> > Lab.   ++39 0116705408
>> > Fax    ++39 0119038639
>> > Mobile ++39 3333827080
>> > email: raffaele.calogero at unito.it
>> >        raffaele[dot]calogero[at]gmail[dot]com
>> > www:   http://www.bioinformatica.unito.it
>> > Info: http://publicationslist.org/raffaele.calogero
>> >
>> >
>> >       [[alternative HTML version deleted]]
>> >
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>> 
>> ------------------------------
>> Mark Robinson, PhD (Melb)
>> Epigenetics Laboratory, Garvan
>> Bioinformatics Division, WEHI
>> e: m.robinson at garvan.org.au
>> e: mrobinson at wehi.edu.au
>> p: +61 (0)3 9345 2628
>> f: +61 (0)3 9347 0852
>> ------------------------------
>> 
>> 
>> 
>> 
>> 
>> 
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> 
> 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
> 

------------------------------
Mark Robinson, PhD (Melb)
Epigenetics Laboratory, Garvan
Bioinformatics Division, WEHI
e: m.robinson at garvan.org.au
e: mrobinson at wehi.edu.au
p: +61 (0)3 9345 2628
f: +61 (0)3 9347 0852
------------------------------






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The information in this email is confidential and intend...{{dropped:6}}