[BioC] help with limma contrast matrix

[Kimpel, Mark W]

I would appreciate advice on how to construct a contrast matrix for a
5X2 ANOVA design. Briefly, I have a genomic experiment to analyze that
compares 5 brain regions in 2 strains of rats. We are interested in
discovering overall differences between strains (collapsing all brain
regions together) but also discovering differences that may only be
expressed in one brain region.

I have attempted to construct the appropriate matrix with the code
listed below, but it does not work. I seem to get differences between
strains, but all the brain region contrasts give exactly the same
results, so I know something isn't correct.

contrast <-makeContrasts(

    (
        (NPAccumbens  + NPAmygdala + NPHippocampus + NPPrefrontal_Cortex
+ NPStriatum) -    #all regions of strain "NP"
        
        (PAccumbens  + PAmygdala + PHippocampus + PPrefrontal_Cortex +
PStriatum)           #all regions of strain "P"
        
    ),
    
    (NPAccumbens - PAccumbens),                    
		#accumbens region of both strains
        
    (NPAmygdala - PAmygdala),                     
		#amygdala region of both strains
        
    (NPHippocampus - PHippocampus),               
		#hippocampus region of both strains
        
    (NPPrefrontal_Cortex - PPrefrontal_Cortex),    
		#Prefrontal_Cortex region of both strains
        
    (NPStriatum - PStriatum),                        
		#striatum region of both strains
        
    levels=design)


Thanks!

Mark

Mark W. Kimpel MD

 

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