Dear List,
we are working on an RNA-Seq experiment including 3 different cultivars of the same plant species during a drought treatment.
The plants have been grown in two different growth chambers. When analysing the data we had to find out that the "effect" of the chambers on gene expression is actually stronger than the drought treatment effect. Hence, we want to remove this (technical) chamber effect from the analysis in order to extract only those genes that are differentially expressed in response to the drought treatment.
Unfortunately we are a bit stuck with correctly formulating the design matrix needed to accomplish this analysis using edgeR. We created a design matrix without the chamber factor according to the manual like this:
but the situation we are facing looks like this:
How do we correctly set up the design matrix to eliminate the chamber effect and is there a way to explicitly define the contrasts we want to extract (analogous to the contrast matrix function in limma?)
thanks for any help
we are working on an RNA-Seq experiment including 3 different cultivars of the same plant species during a drought treatment.
The plants have been grown in two different growth chambers. When analysing the data we had to find out that the "effect" of the chambers on gene expression is actually stronger than the drought treatment effect. Hence, we want to remove this (technical) chamber effect from the analysis in order to extract only those genes that are differentially expressed in response to the drought treatment.
Unfortunately we are a bit stuck with correctly formulating the design matrix needed to accomplish this analysis using edgeR. We created a design matrix without the chamber factor according to the manual like this:
Code:
genotypes <- factor( c("cultivar1", "cultivar1", "cultivar1", "cultivar1", "cultivar2", "cultivar2", "cultivar2", "cultivar2") ) groups <- factor( c("drought", "drought", "watered", "watered", "drought", "drought", "watered", "watered") ) design <- model.matrix(~genotypes + groups) view(design) (Intercept) genotypes groups 1 1 1 0 2 1 1 0 3 1 1 1 4 1 1 1 5 1 0 0 6 1 0 0 7 1 0 1 8 1 0 1
Code:
1 cultivar1 drought chamber1 2 cultivar1 drought chamber2 3 cultivar1 watered chamber1 4 cultivar1 watered chamber2 5 cultivar2 drought chamber1 6 cultivar2 drought chamber2 7 cultivar2 watered chamber1 8 cultivar2 watered chamber2
thanks for any help
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