Dear all,


I have a question regarding the way to properly design biological replicate and technical replicates.

In the projet, we have two groups of sample : R (respond to the treatment), NR (no response).
For each groups, there is several replicates (3) that come from different patients (so not strictely biological).
Each patient have two sample : one at day one(d1), the second after one week (d7)
The hybridization are planned to be performed in dual color, with dye-sap.

Here is the target file :

Cy3	Cy5	Patient	Issu	TechnicalReplicat
d1	d7	A	R	1
d7	d1	A	R	1
d1	d7	B	R	2
d7	d1	B	R	2
d1	d7	C	R	3
d7	d1	C	R	3
d1	d7	D	NR	4
d7	d1	D	NR	4
d1	d7	E	NR	5
d7	d1	E	NR	5
d1	d7	F	NR	6
d7	d1	F	NR	6


my question is how to properly design the matrix design and the contrast matrix to answer the difference between the two groups ?
Which column should I use as biological replicat (so as I can use the duplicateCorrelation function) ?



here is how I've started, but I can't figure out how to use duplicateCorrelation with this design

design <- cbind(
		R_D7vsD1  = c(-1,1,-1,1,-1,1,  0,0, 0,0, 0,0),
		NR_D7vsD1 = c( 0,0,0,0,0,0,-1,1,-1,1,-1,1)
)
fit <- lmFit(MAn, design, weights = NULL)
cont.mat = makeContrasts(PDvsSD = PD_TvsBL - SD_TvsBL, levels = design)
fit2 <- contrasts.fit(fit, cont.mat)
fit2 <- eBayes(fit2)




Many thanks by advance for any help.

--
Guillaume
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