Entering edit mode
Dario Beraldi
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20
@dario-beraldi-4593
Last seen 9.6 years ago
Hello,
I'm using limma to analyze affymetrix arrays from a time course
experiment (time points 0, 2, 7, 24). We are interested in comparing 0
vs 2, 0 vs 7, 0 vs 24.
The experimental design looks like this:
anim_id<- as.factor(rep(c('L1', 'L2', 'L3'), each= 4))
time_point<- rep(c(0, 2, 7, 24), 3)
slide_id<- paste(anim_id, time_point, sep= '.')
(targets<- data.frame(slide_id, anim_id, time_point))
# slide_id anim_id time_point
#1 L1.0 L1 0
#2 L1.2 L1 2
#3 L1.7 L1 7
#4 L1.24 L1 24
#5 L2.0 L2 0
#6 L2.2 L2 2
#7 L2.7 L2 7
#8 L2.24 L2 24
#9 L3.0 L3 0
#10 L3.2 L3 2
#11 L3.7 L3 7
#12 L3.24 L3 24
I wanted to analyze all the arrays at once to fully take advantage of
the eBayes function. Also I wanted to capitalize on the fact that the
same animal is sampled at each time point (0, 2, 7, 24 hours).
So I set up the design matrix like this:
design<- model.matrix(~0+anim_id)
rownames(design)<- paste(targets$anim_id, targets$time_point, sep=
'.')
design<- cbind(design, tp.2vs0= c(0,1,0,0, 0,1,0,0, 0,1,0,0)) ## 0 vs
2
design<- cbind(design, tp.7vs0= c(0,0,1,0, 0,0,1,0, 0,0,1,0)) ## 0 vs
7
design<- cbind(design, tp.24vs0= c(0,0,0,1, 0,0,0,1, 0,0,0,1)) ## 0 vs
24
design
# anim_idL1 anim_idL2 anim_idL3 tp.2vs0 tp.7vs0 tp.24vs0
# L1.0 1 0 0 0 0 0
# L1.2 1 0 0 1 0 0
# L1.7 1 0 0 0 1 0
# L1.24 1 0 0 0 0 1
# L2.0 0 1 0 0 0 0
# L2.2 0 1 0 1 0 0
# L2.7 0 1 0 0 1 0
# L2.24 0 1 0 0 0 1
# L3.0 0 0 1 0 0 0
# L3.2 0 0 1 1 0 0
# L3.7 0 0 1 0 1 0
# L3.24 0 0 1 0 0 1
Now I can procede 'as usual':
fit <- lmFit(limma_affy, design)
fit <- eBayes(fit)
topTable(fit, coef= 4) ## 0 vs 2
topTable(fit, coef= 5) ## 0 vs 7
topTable(fit, coef= 6) ## 0 vs 24
I verified that the logFC I get from topTable is the same as the logFC
calculated manually, which tells me that the design is correct.
However, I would like to make sure that this procedure is correct,
Would anyone be so kind to confirm it?
All the best
Dario
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