[BioC] Limma-design matrix for technical replication

Jenny Drnevich drnevich at illinois.edu
Tue Aug 12 17:28:15 CEST 2008

Hi Katerina,

There is an example of how to deal with both biological and technical 
replicates in the limma vignette, section 8.2. There is even an 
example for dye-swaps, which I believe fits your design exactly. To 
get the limma vignette:

 > limmaUsersGuide()

Good luck,

At 10:06 AM 8/11/2008, =?iso-8859-2?Q?Kate=F8ina_Kepkov=E1?= wrote:
>Dear all,
>As a complete newbie to microarrays, I am trying to analyze experiment with
>following design: Two samples (differentiated versus undifferentiated cells)
>were compared directly on two-color oligo array, with 3 biological
>replicates (different cell sources) and 4 technical replicates (arrays) per
>biological replicate (12 arrays altogether). In every set of technical
>replicates two arrays are dye-swap. I am not sure how to handle the
>technical and biological replication when trying to fit linear model. We are
>interested just in overall comparison differentiated versus undifferentiated
>I have arrived to following setup:
>Targets file is:
>SlideNumber     FileName        Cy3     Cy5
>1       1.gpr   N1      D1
>2       2.gpr   N1      D1
>3       3.gpr   D1      N1
>4       4.gpr   D1      N1
>5       5.gpr   N2      D2
>6       6.gpr   N2      D2
>7       7.gpr   D2      N2
>8       8.gpr   D2      N2
>9       9.gpr   N3      D3
>10      10.gpr  N3      D3
>11      11.gpr  D3      N3
>12      12.gpr  D3      N3
>Where N means undifferentiated and D differentiated cells and 1-3 are
>biological replicates.
>Is the following design correct one? Or is there a better way to obtain
>relevant information?
>Is this extensible for more/less biological replicates?
>design <- cbind(D1vsN1 = c(1,1,-1,-1,0,0,0,0,0,0,0,0), D2vsN2 =
>c(0,0,0,0,1,1,-1,-1,0,0,0,0), D3vsN3 = c(0,0,0,0,0,0,0,0,1,1,-1,-1))
>fit <- lmFit(MA, design)
>cont.matrix <- makeContrasts(DvsN = (D1vsN1 + D2vsN2 + D3vsN3)/3, levels =
>fit2 <- contrasts.fit(fit, cont.matrix)
>fit2 <- eBayes(fit2)
>Sorry if I am asking something obvious and thank you in advance for your
>Best regards,
>Katerina Kepkova
>Laboratory of developmental biology
>Department of Reproductive and Developmental Biology
>Institute of Animal Physiology and Genetics of the AS  CR, v.v.i.
>Rumburska 89, Libechov 277 21
>Czech Republic
>tel:     +420 315 639 534
>fax:     +420 315 639 510
>e-mail: kepkova at iapg.cas.cz
>Bioconductor mailing list
>Bioconductor at stat.math.ethz.ch
>Search the archives: 

Jenny Drnevich, Ph.D.

Functional Genomics Bioinformatics Specialist
W.M. Keck Center for Comparative and Functional Genomics
Roy J. Carver Biotechnology Center
University of Illinois, Urbana-Champaign

330 ERML
1201 W. Gregory Dr.
Urbana, IL 61801

ph: 217-244-7355
fax: 217-265-5066
e-mail: drnevich at illinois.edu

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