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
>
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