[BioC] limma lmFit error: Coefficients not estimable

[Freydin, Maxim]

Hello,

I found that the question was already answered, but the solution proposed did not work for me.

Could you please help to resolve an issue with limma I faced. Please see the attachment. I have a set of arrays for 4 groups of patients (Atype) taken at two time points (Visit). Every patient was analyzed twice, so they are paired and I cannot ignore this. The questions are 1) what genes are differentially expressed between time points in every group; 2) what genes are differentially expressed between disease types at V1 and V3.
I make a design matrix this way:

atype<-factor(c("lco","bri","bri","bri","con","con","con","lco","mild","mild","mild","lco","con","con","lco","bri","bri","bri","con","con","con","lco","mild","mild","mild","lco","con","con"))
patient<-factor(c("x1","x8","x10","x14","x17","x18","x19","x20","x24","x26","x27","x29","x31","x38","x1","x8","x10","x14","x17","x18","x19","x20","x24","x26","x27","x29","x31","x38"))
visit<-factor(c("v1","v1","v1","v1","v1","v1","v1","v1","v1","v1","v1","v1","v1","v1","v3","v3","v3","v3","v3","v3","v3","v3","v3","v3","v3","v3","v3","v3"))
fact<-paste(visit,atype,sep=".")
fact<-as.factor(fact)
design<-model.matrix(~0+fact+patient)
colnames(design)<-c(levels(fact),colnames(design[,9:21]))

cont<-makeContrasts(Mild.Bri.V1=v1.mild-v1.bri,Mild.Bri.V3=v3.mild-v3.bri,Mild.Bri.V1.V3=(v1.mild-v3.mild)-(v1.bri-v3.bri),Mild.Con.V1=v1.mild-v1.con,Mild.Con.V3=v3.mild-v3.con,Mild.Con.V1.V3=(v1.mild-v3.mild)-(v1.con-v3.con),Mild.Lco.V1=v1.mild-v1.lco,Mild.Lco.V3=v3.mild-v3.lco,Mild.Lco.V1.V3=(v1.mild-v3.mild)-(v1.lco-v3.lco),Bri.Con.V1=v1.bri-v1.con,Bri.Con.V3=v3.bri-v3.con,Bri.Con.V1.V3=(v1.bri-v3.bri)-(v1.con-v3.con),Bri.Lco.V1=v1.bri-v1.lco,Bri.Lco.V3=v3.bri-v3.lco,Bri.Lco.V1.V3=(v1.bri-v3.bri)-(v1.lco-v3.lco),Lco.Con.V1=v1.lco-v1.con,Lco.Con.V3=v3.lco-v3.con,Lco.Con.V1.V3=(v1.lco-v3.lco)-(v1.con-v3.con),levels=(design))

fit<-lmFit(eset.norm,design)
  # Coefficients not estimable: patientx27 patientx38 patientx8

fit2<-eBayes(contrasts.fit(fit,cont))


In my opinion, the design matrix looks all right, so does it mean there is a bug in limma?
Thanks

Best wishes,
Maxim
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