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 [[alternative HTML version deleted]]