I am trying to follow the "Load Data" Vignette in the section entitled "quality control through data exploration". I am using R1.7.1 and updated Bioconductor Tuesday (yesterday) on a Windows XP machine. I have 23 U133A chips that I am trying to examine. As I followed the process I was able to do all the actions (histogram, boxplot, RNA degradation plot) on my dataset. > data<-ReadAffy() > data AffyBatch object size of arrays=712x712 features (91097 kb) cdf=HG-U133A (22283 affyids) number of samples=23 number of genes=22283 annotation=hgu133a > I then wanted to see what my data looked like following normalization (similar to what is described from pg 23 on). However, due to memeory issues I could not run the example in a manner similar to that described > normalized.data<-normalize(data[1:23]) Error: cannot allocate vector of size 91091 Kb so I ran justRMA > normalized.data<-justRMA() Background correcting Normalizing Calculating Expression > sampleNames(normalized.data) [1] "1082_U133A.CEL" "1083_HG_U133A.CEL" "1085_U133A.CEL" [4] "1087_U133A.CEL" "109335_U133A.CEL" "109338_U133A.CEL" [7] "109341_U133A.CEL" "109404_HG_U133A.CEL" "1095_U133A.CEL" [10] "110256_U133A.CEL" "110259_U133A.CEL" "110442_HG_U133A.CEL" [13] "110443_HG_U133A.CEL" "110444_HG_U133A.CEL" "110445_HG_U133A.CEL" [16] "1349_U133A.CEL" "1460_U133A.CEL" "7021.31_HG_U133A.CEL" [19] "7113.3_HG_U133A.CEL" "7118.3_HG_U133A.CEL" "7149.3_HG_U133A.CEL" [22] "813.31_HG_U133A.CEL" "F1089_U133A.CEL" > normalized.data Expression Set (exprSet) with 22283 genes 23 samples phenoData object with 1 variables and 23 cases varLabels sample: arbitrary numbering > Now I can not get boxplot or histogram. I have not tried RNA degradation plot. > boxplot(normalized.data,col=c(2,2,3,3)) Error in boxplot.default(normalized.data, col = c(2, 2, 3, 3)) : invalid first argument > > hist(normalized.data[1:2]) Error in hist.default(normalized.data[1:2]) : `x' must be numeric > If Ihave created an incompatible objecct with justRMA, how do I create the correct object type? If not, what suggestions do you have. Michael Barnes, Ph.D.

> > boxplot(normalized.data,col=c(2,2,3,3)) > Error in boxplot.default(normalized.data, col = c(2, 2, 3, 3)) : > invalid first argument > > > > > hist(normalized.data[1:2]) > Error in hist.default(normalized.data[1:2]) : > `x' must be numeric > > > > If Ihave created an incompatible objecct with justRMA, how do I create > the correct object type? If not, what suggestions do you have. I think the problem may be that you are passing the whole expression set, rather than just the expression levels themselves...try hist(normalized.data@exprs[,1:2]) # should give you the first two samples hist(normalized.data@exprs) # should give you a histogram of all expression levels similarly for the boxplot, however if you change the matrix to a data frame, it will plot each chip seperately boxplot(as.data.frame(normalized.data@exprs),col=c(2,2,3,3)) I am not sure the RNA degradation plot will work on normalised values hope this helps claire -------------------------------------------------------- This email is confidential and intended solely for the use o...{{dropped}}

The hist and boxplot functions you are trying to use are intended for AffyBatch objects, not exprSet objects, which is what you have after running RMA. Also, I don't think you will be able to simply normalize an AffyBatch if you use the default quantile normalization. I reported a bug in normalize.AffyBatch.quantiles about two months ago, where I couldn't normalize 9 U-133A chips with 2 Gb RAM. To my knowledge this has not been fixed. You might try normalize(abatch, method="invariantset") or one of the other normalization procedures. HTH, Jim James W. MacDonald Affymetrix and cDNA Microarray Core University of Michigan Cancer Center 1500 E. Medical Center Drive 7410 CCGC Ann Arbor MI 48109 734-647-5623 >>> "Michael Barnes" <michael.barnes@cchmc.org> 08/20/03 11:23AM >>> I am trying to follow the "Load Data" Vignette in the section entitled "quality control through data exploration". I am using R1.7.1 and updated Bioconductor Tuesday (yesterday) on a Windows XP machine. I have 23 U133A chips that I am trying to examine. As I followed the process I was able to do all the actions (histogram, boxplot, RNA degradation plot) on my dataset. > data<-ReadAffy() > data AffyBatch object size of arrays=712x712 features (91097 kb) cdf=HG-U133A (22283 affyids) number of samples=23 number of genes=22283 annotation=hgu133a > I then wanted to see what my data looked like following normalization (similar to what is described from pg 23 on). However, due to memeory issues I could not run the example in a manner similar to that described > normalized.data<-normalize(data[1:23]) Error: cannot allocate vector of size 91091 Kb so I ran justRMA > normalized.data<-justRMA() Background correcting Normalizing Calculating Expression > sampleNames(normalized.data) [1] "1082_U133A.CEL" "1083_HG_U133A.CEL" "1085_U133A.CEL" [4] "1087_U133A.CEL" "109335_U133A.CEL" "109338_U133A.CEL" [7] "109341_U133A.CEL" "109404_HG_U133A.CEL" "1095_U133A.CEL" [10] "110256_U133A.CEL" "110259_U133A.CEL" "110442_HG_U133A.CEL" [13] "110443_HG_U133A.CEL" "110444_HG_U133A.CEL" "110445_HG_U133A.CEL" [16] "1349_U133A.CEL" "1460_U133A.CEL" "7021.31_HG_U133A.CEL" [19] "7113.3_HG_U133A.CEL" "7118.3_HG_U133A.CEL" "7149.3_HG_U133A.CEL" [22] "813.31_HG_U133A.CEL" "F1089_U133A.CEL" > normalized.data Expression Set (exprSet) with 22283 genes 23 samples phenoData object with 1 variables and 23 cases varLabels sample: arbitrary numbering > Now I can not get boxplot or histogram. I have not tried RNA degradation plot. > boxplot(normalized.data,col=c(2,2,3,3)) Error in boxplot.default(normalized.data, col = c(2, 2, 3, 3)) : invalid first argument > > hist(normalized.data[1:2]) Error in hist.default(normalized.data[1:2]) : `x' must be numeric > If Ihave created an incompatible objecct with justRMA, how do I create the correct object type? If not, what suggestions do you have. Michael Barnes, Ph.D. _______________________________________________ Bioconductor mailing list Bioconductor@stat.math.ethz.ch https://www.stat.math.ethz.ch/mailman/listinfo/bioconductor

Thanks for the reply. Is it possible to do similar opperations on my data (get justRMA values) and look at them in a similar way to the probelevel data? I liked the Vignette and how it showed the plots for the probelevel data, then used "normalize" and then looked at the plots again. Does normalize return probe level data again? If not, do you have a suggestion how I might accomplishe this? The memory comment was simply to explain what had happened. I was one of those guided through the memeory issues which is why I chose justRMA instead of "normalize" when that failed. Michael Barnes, Ph.D. >>> ririzarr@jhsph.edu 08/20/03 12:01PM >>> be aware that ReadAffy gives you probe level data. justRMA gives you gene expression level data. i prefer doing QC on the probe level becuase, for one, the spatial proprerties are preserved. regarding the memory problem there have been many posts on the list. look through the mail archive and/or look at the R Windows FAQ. On Wed, 20 Aug 2003, Michael Barnes wrote: > I am trying to follow the "Load Data" Vignette in the section entitled > "quality control through data exploration". I am using R1.7.1 and > updated Bioconductor Tuesday (yesterday) on a Windows XP machine. I > have 23 U133A chips that I am trying to examine. > > As I followed the process I was able to do all the actions (histogram, > boxplot, RNA degradation plot) on my dataset. > > data<-ReadAffy() > > > data > AffyBatch object > size of arrays=712x712 features (91097 kb) > cdf=HG-U133A (22283 affyids) > number of samples=23 > number of genes=22283 > annotation=hgu133a > > > > I then wanted to see what my data looked like following normalization > (similar to what is described from pg 23 on). However, due to memeory > issues I could not run the example in a manner similar to that > described > > > normalized.data<-normalize(data[1:23]) > Error: cannot allocate vector of size 91091 Kb > > so I ran justRMA > > > normalized.data<-justRMA() > Background correcting > Normalizing > Calculating Expression > > > sampleNames(normalized.data) > [1] "1082_U133A.CEL" "1083_HG_U133A.CEL" "1085_U133A.CEL" > > [4] "1087_U133A.CEL" "109335_U133A.CEL" "109338_U133A.CEL" > > [7] "109341_U133A.CEL" "109404_HG_U133A.CEL" "1095_U133A.CEL" > > [10] "110256_U133A.CEL" "110259_U133A.CEL" > "110442_HG_U133A.CEL" > [13] "110443_HG_U133A.CEL" "110444_HG_U133A.CEL" > "110445_HG_U133A.CEL" > [16] "1349_U133A.CEL" "1460_U133A.CEL" > "7021.31_HG_U133A.CEL" > [19] "7113.3_HG_U133A.CEL" "7118.3_HG_U133A.CEL" > "7149.3_HG_U133A.CEL" > [22] "813.31_HG_U133A.CEL" "F1089_U133A.CEL" > > > normalized.data > Expression Set (exprSet) with > 22283 genes > 23 samples > phenoData object with 1 variables and 23 cases > varLabels > sample: arbitrary numbering > > > > Now I can not get boxplot or histogram. I have not tried RNA > degradation plot. > > > boxplot(normalized.data,col=c(2,2,3,3)) > Error in boxplot.default(normalized.data, col = c(2, 2, 3, 3)) : > invalid first argument > > > > > hist(normalized.data[1:2]) > Error in hist.default(normalized.data[1:2]) : > `x' must be numeric > > > > If Ihave created an incompatible objecct with justRMA, how do I create > the correct object type? If not, what suggestions do you have. > > Michael Barnes, Ph.D. > > _______________________________________________ > Bioconductor mailing list > Bioconductor@stat.math.ethz.ch > https://www.stat.math.ethz.ch/mailman/listinfo/bioconductor >