MA Plots after Normalization ? We have tried limma too !

Yes..!!! We got a NEW problem!
We use maNorm() to normalize the dataset and want to compare the MA plots before/after normalization. However when we will apply maQualityPlots, it showed error message again!

>NormalizedData=maNorm(rawdata) >maQualityPlots(NormalizedData, dev="JPEG") Error in 1:dim(mrawObj)[2] : NA/NaN argument > summary(NormalizedData) Normalized intensity data: Object of class marrayNorm. . . > NormalizedData An object of class "marrayNorm" @maA 6Hs.195.1.gpr 6Hs.168.gpr 6Hs.166.gpr 6Hs.187.1.gpr 6Hs.194.gpr 6Hs.243.1.gpr [1,] 6.895683 7.298095 NA NA NA 3.336213 [2,] 5.965369 6.322153 5.608070 3.953445 NA 5.213656 [3,] 6.332001 5.879320 6.272724 3.390680 6.759634 5.960176 . . . >write.marray(NormalizedData) NULL

1) maNorm() to normalize the dataset 2) tried to get MA plots 3) check the normalized dataset -> Yes, it's normailized 4) By looking the data, we found the NA data within normalized dataset. We guess that's the reason made maQualityPlots() not work! 5) But we still can write the normalized data into a new file by using write.marray().

We decided to try another package for normalization, which is limma

>source("http://bioconductor.org/biocLite.R") >biocLite("limma") >library(limma) >target <- readTargets("TargetBeta7.txt") >f <- function(x) as.numeric(x$Flags > -99) >RG<-read.maimages(target,source="genepix",wt.fun=f) Read 6Hs.195.1.gpr Read 6Hs.168.gpr Read 6Hs.166.gpr Read 6Hs.187.1.gpr Read 6Hs.194.gpr Read 6Hs.243.1.gpr >RG<-backgroundCorrect(RG, method="normexp",offset=50) Corrected array 1 Corrected array 2 Corrected array 3 Corrected array 4 Corrected array 5 Corrected array 6 >maQualityPlots(RG, dev="jpeg") >nRG<-normalizeWithinArrays(RG) >maQualityPlots(nRG, dev="jpeg") Error in do.call("gpFlagWt", list(mraw@maW)) : no slot of name "maW" for this object of class "MAList"

1) Install it first 2) then load the package 3) read in the list for *.gpr file 4) Set up a filter so that any spot with a flag of −99 or less gets zero weight (based on limma userguide) 5) Read in the Red/Green raw intensities to "RG" 6) Do an adaptive background correction which is optional but recommended for GenePix data 7) get the MA plots 8) run normalization 9) try to get MA plots after normalization -> failed again