[BioC] edgeR - paired samples with multifactorial design - errors

[James W. MacDonald]

Hi Preethy,

This experiment is very similar to the example in part 3.5 of the edgeR 
User's guide, starting on page 31.

Best,

Jim


On 4/25/2014 11:29 AM, Preethy Venkat Ram wrote:
> Hi Devon,
>
> Thanks for the replies both on biostars and here.  Sorry for
> crossposting.  Both
> mailing lists and discussion have been very helpful to me.
>
> But, I rarely see replies from BioC package maintainers at biostars.
>
> All the samples are paired and I have them all correct - I mean none of
> them are empty
> I tried different designs. But ending up with the same error message.
> What I want to do: Getting DE genes between
> (Phenotype1.Before-Phenotype1.After) & (Phenotype2.Before-Phenotype2.After)
>
> pdata here:
>
> pair=rep(c(1:45), each=2)
> Treat=rep(c("Before", "After"),45)
> Phenotype=rep(c(1, 1, 2, 2, 1, 1, 1, 2, 1, 1, 1, 2, 1,  2, 2, 2, 2, 2, 2,
> 2, 1, 1, 1, 1, 1, 1, 2, 2, 1,  2, 2, 2, 2, 2, 2, 2, 2, 1, 2, 1, 2, 2, 1, 2,
> 2), each=2)
> pdata<-data.frame(pair,Treat,Phenotype)
>
>
> Preethy
>
>
>
> On Fri, Apr 25, 2014 at 4:53 PM, Devon Ryan <dpryan at dpryan.com> wrote:
>
>> Hi Preethy,
>>
>> You likely want:
>>
>> design=model.matrix(~pair+Treat:Phenotype, data=pdata)
>>
>> If that still yields the error, then you'll need to share "pdata" or
>> "design". Also, please don't crosspost on both this list and biostars (
>> https://www.biostars.org/p/98907/), it duplicates the community effort.
>>
>> Devon
>>
>>
>> --
>> Devon Ryan, Ph.D.
>> Email: dpryan at dpryan.com
>> Laboratory for Molecular and Cellular Cognition
>> German Centre for Neurodegenerative Diseases (DZNE)
>> Ludwig-Erhard-Allee 2
>> 53175 Bonn
>> Germany
>> <devon.ryan at dzne.de>
>>
>>
>> On Fri, Apr 25, 2014 at 11:15 AM, Preethy [guest] <guest at bioconductor.org>wrote:
>>
>>> Hi All,
>>>
>>> I had been trying to do DE analysis of my RNAseq experiment using edgeR
>>> and am having some isssues. The details of the Experiment and the R code I
>>> tried below:
>>>
>>> (a) Paired experimental design with 45 pairs
>>> (b) Treatment: "Before" and "After"
>>> (c) Phenotype: 1 & 2
>>> Aim: Look for DE genes between Phenotype 1 and 2 upon treatment taking
>>> into account the paired design
>>>
>>> The R code tried:
>>>
>>> library(edgeR)
>>> counts<-read.delim(file="counts.dat",header=T)
>>> pair=factor(pdata$pair)
>>> Treat=factor( pdata$treat)
>>> Phenotype=factor(pdata$pheno)
>>> group<-paste(Treat,Phenotype,sep=".")
>>> design=model.matrix(~pair+Treat:Phenotype, data=counts)
>>> counts.DGEList<-DGEList(counts, group=group)
>>> y<-calcNormFactors(counts.DGEList)
>>> y<-estimateCommonDisp(y, design)
>>> y<-estimateGLMTrendedDisp(y, design)
>>>
>>>
>>> Error message I get:
>>>
>>> Error in glmFit.default(y, design = design, dispersion = dispersion,
>>> offset = offset,  :
>>>    Design matrix not of full rank.  The following coefficients not
>>> estimable:
>>>   TreatBefore:Phenotype1 TreatBefore:Phenotype2
>>>
>>>
>>> Any idea to solve this out?
>>>
>>> Thanks,
>>> Preethy
>>>
>>>   -- output of sessionInfo():
>>>
>>> R version 3.1.0 (2014-04-10)
>>> Platform: x86_64-pc-linux-gnu (64-bit)
>>>
>>> locale:
>>>   [1] LC_CTYPE=fi_FI.UTF-8       LC_NUMERIC=C
>>>   [3] LC_TIME=en_GB.UTF-8        LC_COLLATE=en_GB.UTF-8
>>>   [5] LC_MONETARY=fi_FI.UTF-8    LC_MESSAGES=en_GB.UTF-8
>>>   [7] LC_PAPER=en_GB.UTF-8       LC_NAME=C
>>>   [9] LC_ADDRESS=C               LC_TELEPHONE=C
>>> [11] LC_MEASUREMENT=en_GB.UTF-8 LC_IDENTIFICATION=C
>>>
>>> attached base packages:
>>> [1] stats     graphics  grDevices utils     datasets  methods   base
>>>
>>> other attached packages:
>>> [1] edgeR_3.4.2   limma_3.18.13
>>>
>>>
>>> --
>>> Sent via the guest posting facility at bioconductor.org.
>>>
>>> _______________________________________________
>>> Bioconductor mailing list
>>> Bioconductor at r-project.org
>>> https://stat.ethz.ch/mailman/listinfo/bioconductor
>>> Search the archives:
>>> http://news.gmane.org/gmane.science.biology.informatics.conductor
>>>
>>
> 	[[alternative HTML version deleted]]
>
> _______________________________________________
> Bioconductor mailing list
> Bioconductor at r-project.org
> https://stat.ethz.ch/mailman/listinfo/bioconductor
> Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor

-- 
James W. MacDonald, M.S.
Biostatistician
University of Washington
Environmental and Occupational Health Sciences
4225 Roosevelt Way NE, # 100
Seattle WA 98105-6099