
Thank you

Ugo


From: Michael Love <michaelisaiahlove@gmail.com>
Date: Wed, 20 Nov 2013 10:21:10 -0500
To: Ugo Borello <ugo.borello@inserm.fr>
Cc: "bioconductor@r-project.org" <bioconductor@r-project.org>
Subject: Re: [BioC] DESeq2 multifactorial formula

hi Ugo,

The problem with the experimental design is that all the males are in batch
3, so you can't separate the effect of these two. There are many ways to
name this problem: linearly dependent covariates, rank deficient design
matrix, etc. My suggestion would be to remove the sex variable. This would
then control for differences due to batch (and in the case of batch 3,
absorbing the male effect). ?You cannot test for significance of the male
effect anyway, because you wouldn't be able to tell apart the male effect
from the batch 3 effect.

hope this helps,

Mike



On Wed, Nov 20, 2013 at 10:03 AM, Ugo Borello <ugo.borello@inserm.fr> wrote:
> Good morning,
> 
> I am trying to run DESeq2 with this design formula
> design <- (~batch+sex+tissue)
> 
> This is what I do from a count matrix:
>> >library(DESeq2)
> 
>> >countTable<- read.table('matrix.txt', header=TRUE, row.names=1)
> 
> 
>> >ConDesign<- data.frame(row.names = colnames(countTable),
> ? ? ? ? ? ? ? ? ? ? ? ?batch = factor(c("1", "2", "3", "1", "2", "3")),
> ? ? ? ? ? ? ? ? ? ? ? ?sex = factor(c("F", "F", "M", "F", "F", "M")),
> ? ? ? ? ? ? ? ? ? ? ? ?tissue = factor(c("Cx", "Cx", "Cx",
> ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? ? "BrS", "BrS", "BrS")))
> 
>> >ConDesign
> ? ? ? ? ? ? ? ?batch sex ?tissue
> Cx_1 ? ? ? ?1 ? ? ? F ? ? Cx
> Cx_2 ? ? ? ?2 ? ? ? F ? ? Cx
> Cx_3 ? ? ? ?3 ? ? ? M ? ?Cx
> BrS_1 ? ? ? 1 ? ? ? F ? ?BrS
> BrS_2 ? ? ? 2 ? ? ? F ? ?BrS
> BrS_3 ? ? ? 3 ? ? ? M ? BrS
> 
> 
> When I run
>> >xx<- model.matrix(~batch+sex+tissue, ConDesign)
> 
> I get
>> >xx
> ? ? ? ? ? ? ? ? (Intercept) batch2 batch3 sexM tissueBrS
> Cx_1 ? ? ? ? ? ? ? ? ? ? ? 1 ? ? ?0 ? ? ?0 ? ? ? ? ?0 ? ? ? ? ?0
> Cx_2 ? ? ? ? ? ? ? ? ? ? ? 1 ? ? ?1 ? ? ?0 ? ? ? ? ?0 ? ? ? ? ?0
> Cx_3 ? ? ? ? ? ? ? ? ? ? ? 1 ? ? ?0 ? ? ?1 ? ? ? ? ?1 ? ? ? ? ?0
> BrS_1 ? ? ? ? ? ? ? ? ? ? ?1 ? ? ?0 ? ? ?0 ? ? ? ? ?0 ? ? ? ? ?1
> BrS_2 ? ? ? ? ? ? ? ? ? ? ?1 ? ? ?1 ? ? ?0 ? ? ? ? ?0 ? ? ? ? ?1
> BrS_3 ? ? ? ? ? ? ? ? ? ? ?1 ? ? ?0 ? ? ?1 ? ? ? ? ?1 ? ? ? ? ?1
> 
> But when I run:
>> >dse<- DESeqDataSetFromMatrix(countData= countTable,
> ? ? ? ? ? ? ? ? ? ? ? ? ? ? ?colData = ConDesign,
> ? ? ? ? ? ? ? ? ? ? ? ? ? ? ?design = ?(~batch+sex+tissue))
> 
> 
> 
> I get this error message:
> Error in validObject(.Object) :
> ? invalid class ³DESeqDataSet² object: the model matrix is not full rank,
> i.e. one or more variables in the design formula are linear combinations of
> the others
> 
> Where is my mistake?
> 
> Thank you for your help,
> Ugo
> 
> 
>> > sessionInfo()
> R version 3.0.1 (2013-05-16)
> Platform: x86_64-apple-darwin10.8.0 (64-bit)
> 
> locale:
> [1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
> 
> attached base packages:
> [1] parallel ?stats ? ? graphics ?grDevices utils ? ? datasets ?methods
> base
> 
> other attached packages:
> ?[1] org.Hs.eg.db_2.9.0 ? ? ?RSQLite_0.11.4 ? ? ? ? ?DBI_0.2-7
> AnnotationDbi_1.22.6
> ?[5] DESeq2_1.0.19 ? ? ? ? ? RcppArmadillo_0.3.910.0 Rcpp_0.10.4
> lattice_0.20-23
> ?[9] Biobase_2.20.1 ? ? ? ? ?GenomicRanges_1.12.5 ? ?IRanges_1.18.3
> BiocGenerics_0.6.0
> 
> loaded via a namespace (and not attached):
> ?[1] annotate_1.38.0 ? ?genefilter_1.42.0 ?grid_3.0.1 ? ? ? ? locfit_1.5-9.1
> RColorBrewer_1.0-5
> ?[6] splines_3.0.1 ? ? ?stats4_3.0.1 ? ? ? survival_2.37-4 ? ?tools_3.0.1
> XML_3.95-0.2
> [11] xtable_1.7-1
> 
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