Question: within array normalization for one color agilent array.??
0
9.7 years ago by
neeraj rana100
neeraj rana100 wrote:
hi, In the package Agi4x44PreProcess I took the expression value as *MeanSigna*l, then by the method *BGandNorm,* the background subtraction and the normalization between the array is done.But the array should be normalized within the array also first.This the script what is given in Agi4x44PreProcess for normalization.I didnt find any option for within the array normalization. > ddNORM = BGandNorm(dd, BGmethod = "half", NORMmethod = "quantile", + foreground = "MeanSignal", background = "BGMedianSignal", + offset = 50, makePLOTpre = FALSE, makePLOTpost = FALSE) please help me out how can we do the normalization within the array before between the array normalization for one color agilent array. thank you. [[alternative HTML version deleted]]
modified 9.7 years ago by Sean Davis21k • written 9.7 years ago by neeraj rana100
Answer: within array normalization for one color agilent array.??
0
9.7 years ago by
Sean Davis21k
United States
Sean Davis21k wrote:
On Fri, Feb 19, 2010 at 1:29 AM, neeraj rana <kushrn at="" gmail.com=""> wrote: > hi, > > In the package Agi4x44PreProcess ?I took the expression value as *MeanSigna*l, > then by the method *BGandNorm,* the background subtraction and the > normalization between the array is done.But the array should be normalized > within the array also first.This the script what is given in > Agi4x44PreProcess for normalization.I didnt find any option for within the > array normalization. > >> ddNORM = BGandNorm(dd, BGmethod = "half", NORMmethod = "quantile", > + foreground = "MeanSignal", background = "BGMedianSignal", > + offset = 50, makePLOTpre = FALSE, makePLOTpost = FALSE) > > please help me out how can we do the normalization within the array before > between the array normalization for one color agilent array. For one-color arrays, it usually suffices to perform between-array normalization. Sean
Hi all, I am trying to read and visualize data obtained from an agilent chip. For this purpose I use limma package and the read.maimages function as follows fileRG.raw <- read.maimages("File",source="agilent") FileRG gets read correctly. When I check the dimensions it shows to have 243494 probes which seem to be correctly annotated. I, then attach the layout for the chip from the GAL file: galinfo <- readGAL(fileGAL) fileRG.raw$printer<-getLayout(galinfo) and try to plot R/G according to this prnter layout: imageplot(log2(fileRG.raw$R),fileRG.raw$printer,zlim=c(0,9),main="R-Fo r.RAW") However, I obtain the following error: > Error in imageplot(log2(fileRG.raw$R), fileRG.raw$printer, zlim = c(0, : > Number of image spots does not agree with layout dimensions Which is due to the fact that the gal file has 912*267 dimension corresponding to 243504 probes. Checking the expression raw information I see that there are 10 probes that are tagged as "ignore", which read.maimages ignores convenientl but, then, I can't visualize R and G channels (or MA info when obtained) because of this inconsistency. If I add, by hand, ten probes in the expression file, imageplot works fine but this is not the right thing to do. What is the convenient thing to do? Any suggestion? Thank you in advance Marc -- ----------------------------------------------------- Marc Noguera i Julian, PhD Genomics unit / Bioinformatics Institut de Medicina Preventiva i Personalitzada del C?ncer (IMPPC) B-10 Office Carretera de Can Ruti Cam? de les Escoles s/n 08916 Badalona, Barcelona ADD REPLYlink written 9.7 years ago by Marc Noguera100 Dear Marc, There is a previous answer to another user with the same problem. Source: https://stat.ethz.ch/pipermail/bioconductor/2007-January/015532.html I paste the text here (original by Dr. Gordon K Smyth) ---8<-------------- The error message is telling you that there aren't enough rows of data for the size of the block. The reason is that Agilent doesn't print rows for blank spots and the like, so the block has missing rows, and this causes a problem because the limma functions assume complete blocks. BTW, this is why the function read.maimages() didn't set the$printer information automatically for you. If the file had contained complete data for the block, then it would have. It is possible with a little R programming to expand out your data vector stuff$Rb[,1] to be the right length by adding NAs in the right places, so that you could use imageplot(), but this bit of code is not provided by limma or Bioconductor. Try this: r <- stuff$genes$Row c <- stuff$genes$Col nr <- max(r) nc <- max(c) y <- rep(NA,nr*nc) i <- (r-1)*nc+c y[i] <- log2(stuff$Rb[,1]) imageplot(y,stuff$printer) Best wishes Gordon --->8-------------- I hope that helps, Best, Juan Carlos Oliveros, Ph.D CNB-CSIC, Madrid, Spain Marc Noguera wrote: > Hi all, > I am trying to read and visualize data obtained from an agilent chip. > > For this purpose I use limma package and the read.maimages function as > follows > > fileRG.raw <- read.maimages("File",source="agilent") > > FileRG gets read correctly. When I check the dimensions it shows to have > 243494 probes which seem to be correctly annotated. I, then attach the > layout for the chip from the GAL file: > > galinfo <- readGAL(fileGAL) > fileRG.raw$printer<-getLayout(galinfo) > > and try to plot R/G according to this prnter layout: > > imageplot(log2(fileRG.raw$R),fileRG.raw$printer,zlim=c(0,9),main="R- For.RAW") > > However, I obtain the following error: > > >> Error in imageplot(log2(fileRG.raw$R), fileRG.raw$printer, zlim = c(0, : >> Number of image spots does not agree with layout dimensions >> > Which is due to the fact that the gal file has 912*267 dimension > corresponding to 243504 probes. Checking the expression raw information > I see that there are 10 probes that are tagged as "ignore", which > read.maimages ignores convenientl but, then, I can't visualize R and G > channels (or MA info when obtained) because of this inconsistency. > If I add, by hand, ten probes in the expression file, imageplot works > fine but this is not the right thing to do. > > What is the convenient thing to do? Any suggestion? > > Thank you in advance > > Marc > >