Hi all,
I'm new to DESeq and am having a problem setting up the design matrix for an experiment that has 3 biological conditions with 3 replicates each and an additional factor to take into an experimental batch effect.
I have proceeded in the following manner:
> library(DESeq)
> data <- read.table("aggregated_counts_final.txt", header=T, row.names=1)
> condition <- c(rep("Control",3), rep("Rods",3), rep("Cubes",3))
> batch <- c(rep(c(1,2,3),3))
> Design <- as.data.frame(cbind(condition,batch),row.names=colnames(data))
> CDS <- newCountDataSet(data,Design)
> CDS <- estimateSizeFactors(CDS)
> CDS <- estimateDispersions(CDS)
Error in .local(object, ...) :
None of your conditions is replicated. Use method='blind' to estimate across conditions, or 'pooled-CR', if you have crossed factors.
I can't quite figure out where I've gone wrong... it seems specific to my design matrix as this is very similar to the Pasilla data example, but that works fine on my system. I suspect I'm not understanding something very basic.
My sessionInfo is as follows:
> sessionInfo()
R version 2.15.2 (2012-10-26)
Platform: x86_64-apple-darwin9.8.0/x86_64 (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] stats graphics grDevices utils datasets methods base
other attached packages:
[1] DESeq_1.10.1 lattice_0.20-13 locfit_1.5-8 Biobase_2.18.0 BiocGenerics_0.4.0
loaded via a namespace (and not attached):
[1] annotate_1.36.0 AnnotationDbi_1.20.3 DBI_0.2-5 genefilter_1.40.0 geneplotter_1.36.0 grid_2.15.2 IRanges_1.16.4 parallel_2.15.2
[9] RColorBrewer_1.0-5 RSQLite_0.11.2 splines_2.15.2 stats4_2.15.2 survival_2.37-2 tools_2.15.2 XML_3.95-0.1 xtable_1.7-0
Thank you so much, in advance.
Joshua Pollack
I'm new to DESeq and am having a problem setting up the design matrix for an experiment that has 3 biological conditions with 3 replicates each and an additional factor to take into an experimental batch effect.
I have proceeded in the following manner:
> library(DESeq)
> data <- read.table("aggregated_counts_final.txt", header=T, row.names=1)
> condition <- c(rep("Control",3), rep("Rods",3), rep("Cubes",3))
> batch <- c(rep(c(1,2,3),3))
> Design <- as.data.frame(cbind(condition,batch),row.names=colnames(data))
> CDS <- newCountDataSet(data,Design)
> CDS <- estimateSizeFactors(CDS)
> CDS <- estimateDispersions(CDS)
Error in .local(object, ...) :
None of your conditions is replicated. Use method='blind' to estimate across conditions, or 'pooled-CR', if you have crossed factors.
I can't quite figure out where I've gone wrong... it seems specific to my design matrix as this is very similar to the Pasilla data example, but that works fine on my system. I suspect I'm not understanding something very basic.
My sessionInfo is as follows:
> sessionInfo()
R version 2.15.2 (2012-10-26)
Platform: x86_64-apple-darwin9.8.0/x86_64 (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] stats graphics grDevices utils datasets methods base
other attached packages:
[1] DESeq_1.10.1 lattice_0.20-13 locfit_1.5-8 Biobase_2.18.0 BiocGenerics_0.4.0
loaded via a namespace (and not attached):
[1] annotate_1.36.0 AnnotationDbi_1.20.3 DBI_0.2-5 genefilter_1.40.0 geneplotter_1.36.0 grid_2.15.2 IRanges_1.16.4 parallel_2.15.2
[9] RColorBrewer_1.0-5 RSQLite_0.11.2 splines_2.15.2 stats4_2.15.2 survival_2.37-2 tools_2.15.2 XML_3.95-0.1 xtable_1.7-0
Thank you so much, in advance.
Joshua Pollack