Dear all,

I perform 2 color hybridization, using microRNA and Exiqon slides.
slide design: 4 replicates of 1 array
                   each array consits of 8 subarrays,
                   each subarray of 16 columns and 11 rows
                   each mir has 4 replicates

For analysis of the data I use limma GUI, to obtain the p-values of
differentially expressed mirs I carried out loess normalization
(backgroundsubstraction normexp, offset=10) and subsequently applied the
Linear model fit, which works, as long as I treat  each replicate like a
single mir.
I would like to now also about the correlation of the replicates, so I
entered as replicate number 4 and as distance 21 (according to
"Bioinformatics+ computional statistics using R and Bioconductor" the
distance corresponds to the number of spots that lies betweeen the
replicates).
But I get the following message:

"Error in dim(M)<- c(spacing,ndups, ngroups, nslides)
dims[product 5628] do not match the length of object [5632]"

I rechecked the gal file and recounted on the image, 21 is the correct
number (considering the column-distance, using the number of spots of row
distance results in dims[product 0]).
I read the limma guide and checked also the examples on the limma gui
homepage, but it's always either no replicate or only 2 replicates next to
each other.
Do you know a solution?

Also another question:
IF one day I manage to do the duplicate correlation and given that I would
use weights to identfy features with the flag "bad", how will the software
handle this if one or more replicates are flagged bad? will it exclude the
flagged bad replicate from the correlation?
If yes, is there a way to put a minimum limit of 2 unflagged spots per mir,
so that in case of only 1 unflagged replicate of a mir to exclude this mir
from analysis?

Thank you  a lot in advance!
Greetings, Christine








Dr. Christine Völlenkle, Ph.D.
Research Laboratories-Molecular Cardiology
I.R.C.C.S. Policlinico San Donato
Via R. Morandi, 30
20097  S. Donato M.se (MI) Italy
Phone: +39 02 52774 683 (lab)
          +39 02 52774 533 (office)
Fax:    +39 02 52774 666
email: christine.voellenkle@gmail.com

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