[BioC] assist in constructing a design matrix & contrast matrix for Illumina microarray

[Maria Dolores Serafica]

   Dear Bioconductor Staff,


   I sent this email on Oct 27 & would like to know if it was received and if
   someone can help.


   I am analysing Illumina Beadchips (HumanWG6) for 42 SAMPLES.
   Experimental design: (factorial) time course, samples grown on 4
   media; have zero control sampled at onset only[ one way multigroup
   analysis]; 2-3 reps per treatment.

   I used the lumi script by Pan Du et al (2009) for preprocessing
   and normalisation . This part  was okay. The samples were subsetted
   and are R objects.

   SAMPLE 1 REFERENCE SAMPLE  DAY 0 (triplicate)
   SAMPLE 2 MEDIA A DAY 7 (triplicate)
   SAMPLE 3 MEDIA A DAY 14 (triplicate)
   SAMPLE 4 MEDIA A Day 30 (triplicate)
   SAMPLE 5 MEDIA A DAY60  (singleton)
   SAMPLE 6 MEDIA A DAY90 (replicates)
   SAMPLE 7 MEDIA B DAY 7 (triplicate)
   SAMPLE 8 MEDIA B DAY 14 (triplicate)
   SAMPLE 9 MEDIA B DAY 30 (triplicate)
   SAMPLE 10 MEDIA B DAY60 (singleton
   SAMPLE 11 MEDIA B DAY 90 (TRIPLICATE)
   SAMPLE 12 MEDIA C DAY 7 (REPLICATES)
   SAMPLE 13 MEDIA C DAY 14 (REPLICATES)
   SAMPLE 14 MEDIA C DAY 30 (REPLICATES)
   SAMPLE 15 MEDIA D DAY 7 (REPLICATES)
   SAMPLE 16 MEDIA D DAY 14 (REPLCATES)
   SAMPLE 17 MEDIA D DAY 30 (REPLICATES)
   SAMPLE 18 MEDIA D (DAY 90 (REPLICATES

   This is a differentiation experiment.

    I tried the lumi script using the modified limma (PanDu et al 2009) based
   on the Barnes subset data but the script does not apply to my experimental
   set up.  Can someone assist me in making a design matrix and a contrast
   matrix.

   I am new to R and would like to use limma for identifying differentially
   expressed genes.  I have read the limma users guide(Gordon Smyth), LAB4
   limma 2005, James MacDonald's BioC 2009.

   The contrasts (comparisons) I am looking for:
   a. differential gene expression of each sample WITH the reference
   b. early differentiation genes as affected by media, time and both
   c. late differentiation genes as affected by media or both.


   Sincerely,

   Maria