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]] >> > >> > _______________________________________________ >> > Bioconductor mailing list >> > Bioconductor at stat.math.ethz.ch >> > https://stat.ethz.ch/mailman/listinfo/bioconductor >> > Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor >> >> ------------------------------ >> 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 >> ------------------------------ >> >> >> >> >> >> >> ______________________________________________________________________ >> The information in this email is confidential and intend...{{dropped:6}} >> >> _______________________________________________ >> Bioconductor mailing list >> Bioconductor at stat.math.ethz.ch >> https://stat.ethz.ch/mailman/listinfo/bioconductor >> Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor > > 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 ------------------------------ ______________________________________________________________________ The information in this email is confidential and intend...{{dropped:6}}