Dear Gordon, Thanks for the explanation. I didnt realize robust spline was dependent of print-tips. Sincerely, Priscila On Thursday, March 8, 2012, Gordon K Smyth <smyth@wehi.edu.au> wrote: > Dear Priscila, > > You can't apply robust-spline normalization to an Agilent array. Robust-spline in limma is intended to moderate the loess curves over different print-tip groups. But Agilent arrays don't have print-tips. For an Agilent arrays, only global loess normalization is meaningful, and that does not require the layout. > > Best wishes > Gordon > >> Date: Wed, 7 Mar 2012 09:16:22 -0300 >> From: Priscila Grynberg <priscilag@gmail.com> >> To: <bioconductor@stat.math.ethz.ch> >> Subject: [BioC] RobustSpline normalization method applied to agilent >> data >> >> Dear BioCs, >> >> I received some agilent data to analyse using Limma, and I'm facing a >> problem when I tried to apply robustspline normalization within arrays >> method. >> >> ##Reading the files: >> >> RG = read.maimages(targets,,source="agilent", columns = list(G = >> "gMedianSignal", Gb = "gBGMedianSignal", R = "rMedianSignal", Rb = >> "rBGMedianSignal"), annotation = c("Row", "Col","FeatureNum", >> "ControlType","ProbeName","SystematicName")) >> >> ##Background correction: >> >> RGn <- backgroundCorrect (RG, method="normexp", offset=50) >> >> #Normalization step >> >> MA <- normalizeWithinArrays(RGn, method="robustspline") >> >> Error in normalizeWithinArrays(RGn, method = "robustspline") : Layout >> argument not specified >> >> ##Setting the printer information: >> >> nr <- length(unique(RG$genes$Row)) >> >> nc <- length(unique(RG$genes$Col)) >> >> RG$printer <- list(ngrid.r=1,ngrid.c=1,nspot.r=nr,nspot.c=nc) >> >> ## Trying again >> >> MA <- normalizeWithinArrays(RGn, RG$printer, method="robustspline") >> >> Error in X[O, ] <- ns(A[O], df = df, intercept = TRUE) : number of items to >> replace is not a multiple of replacement length >> >> I don't have the gal file like for this agilent slides. Can this be exactly >> the problem? Do you have any sugestions? >> >> >> Cheers, >> >> Priscila >> >> >> -- >> Priscila Grynberg, D.Sc.- Bioinformatics >> Laboratório de Genética Bioquímica >> Universidade Federal de Minas Gerais >> Tel: +55 31 3409-2628 >> CV: http://lattes.cnpq.br/8808643075395963 > > ______________________________________________________________________ > The information in this email is confidential and inte...{{dropped:12}}