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gravatar for Louisa A Rispoli/AS/EXP/UTIA
10.2 years ago by
Hi all- I am fairly new to bioconductor and am very excited about the example described in the affycoretools vignette since 1) it would make my mentor very happy (almost the exact situation that we have in the data set), 2) seemed like a simple solution to a issue I was not sure how to approach and 3) package seemed straightforward and easy to use. This is probably a stupid question arising from the fact that I have yet to fully understand R and bioconductor but I have tried to spend a better portion of the day trying to figure where I am going wrong and having no luck. I am trying to compare samples (two treatment groups) amplified two different ways and see which differentially expressed genes are shared between the two different methodologies. According to the PCA plot I need to compute the expression values for each methodology separately and then combine the data. I can not seem to create the new expression set properly to perform the limma analysis. I have attached a text file with my R code. If anyone could point me in the right direction (either the mistake I am makeing or even the vignette that I should read closer) I would appreciate the help. Sorry if this seems very silly but we do not have anyone here familiar with Bioconductor to go to for help. Thanks Louisa Sorry required a repost, forgot that attachments were stripped. R version 2.7.0 (2008-04-22) Copyright (C) 2008 The R Foundation for Statistical Computing ISBN 3-900051-07-0 > library(affycoretools) Loading required package: affy Loading required package: Biobase Loading required package: tools Welcome to Bioconductor Vignettes contain introductory material. To view, type 'openVignette()'. To cite Bioconductor, see 'citation("Biobase")' and for packages 'citation(pkgname)'. Loading required package: affyio Loading required package: preprocessCore Loading required package: limma Loading required package: GOstats Loading required package: graph Loading required package: GO.db Loading required package: AnnotationDbi Loading required package: DBI Loading required package: RSQLite Loading required package: annotate Loading required package: xtable Loading required package: RBGL Loading required package: Category Loading required package: genefilter Loading required package: survival Loading required package: splines Loading required package: biomaRt Loading required package: RCurl Attaching package: 'biomaRt' The following object(s) are masked from package:annotate : getGO Loading required package: gcrma Loading required package: matchprobes Loading required package: annaffy Loading required package: KEGG.db Attaching package: 'annaffy' The following object(s) are masked from package:RCurl : getURL Warning messages: 1: package 'DBI' was built under R version 2.7.1 2: package 'RSQLite' was built under R version 2.7.1 3: package 'xtable' was built under R version 2.7.1 > pd <-read.AnnotatedDataFrame("pData.txt", sep="\t", header=TRUE, row.names=1) > dat <- ReadAffy(phenoData= pd) > pData(dat) amp trt PolyC-1.CEL PolyA Ctrl PolyC-2.CEL PolyA Ctrl PolyC-3.CEL PolyA Ctrl PolyC-4.CEL PolyA Ctrl PolyC-5.CEL PolyA Ctrl PolyC-6.CEL PolyA Ctrl PolyC-7.CEL PolyA Ctrl PolyC-8.CEL PolyA Ctrl PolyHS-1.CEL PolyA HS PolyHS-2.CEL PolyA HS PolyHS-3.CEL PolyA HS PolyHS-4.CEL PolyA HS PolyHS-5.CEL PolyA HS PolyHS-6.CEL PolyA HS PolyHS-7.CEL PolyA HS PolyHS-8.CEL PolyA HS WTC-1.CEL WT Ctrl WTC-2.CEL WT Ctrl WTC-3.CEL WT Ctrl WTC-4.CEL WT Ctrl WTC-5.CEL WT Ctrl WTC-6.CEL WT Ctrl WTC-7.CEL WT Ctrl WTC-8.CEL WT Ctrl WTHS-1.CEL WT HS WTHS-2.CEL WT HS WTHS-3.CEL WT HS WTHS-4.CEL WT HS WTHS-5.CEL WT HS WTHS-6.CEL WT HS WTHS-7.CEL WT HS WTHS-8.CEL WT HS > eset <-affystart(groups=rep(1:4, each = 8), groupsnames = unique(paste(pData(pd)[,1],pData(pd)[,2], sep="-")), phenoData = pd) Background correcting Normalizing Calculating Expression > eset1 <-affystart(filenames=list.celfiles()[1:16], plot = FALSE, pca=FALSE) Background correcting Normalizing Calculating Expression > eset2 <-affystart(filenames=list.celfiles()[17:32], plot=FALSE, pca=FALSE) Background correcting Normalizing Calculating Expression > eset <- new("ExpressionSet", exprs=cbind(exprs(eset1), exprs(eset2)), phenoData = pd, annotation=annotation(eset1)) Error in validObject(.Object) : invalid class "ExpressionSet" object: sampleNames differ between assayData and phenoData > eset1 ExpressionSet (storageMode: lockedEnvironment) assayData: 24128 features, 16 samples element names: exprs phenoData sampleNames: PolyC-1.CEL, PolyC-2.CEL, ..., PolyHS-8.CEL (16 total) varLabels and varMetadata description: sample: arbitrary numbering featureData featureNames: AFFX-BioB-3_at, AFFX-BioB-5_at, ..., BtAffx.29968.1.S1_at (24128 total) fvarLabels and fvarMetadata description: none experimentData: use 'experimentData(object)' Annotation: bovine > eset2 ExpressionSet (storageMode: lockedEnvironment) assayData: 24128 features, 16 samples element names: exprs phenoData sampleNames: WTC-1.CEL, WTC-2.CEL, ..., WTHS-8.CEL (16 total) varLabels and varMetadata description: sample: arbitrary numbering featureData featureNames: AFFX-BioB-3_at, AFFX-BioB-5_at, ..., BtAffx.29968.1.S1_at (24128 total) fvarLabels and fvarMetadata description: none experimentData: use 'experimentData(object)' Annotation: bovine "If we knew what we were doing, it wouldn't be called Research." - Albert Einstein Louisa Rispoli, Ph.D. Reproductive Physiology Department of Animal Science University of Tennessee, Knoxville A105 Johnson Animal Research and Teaching Unit 1750 Alcoa Highway Knoxville, TN 37920 phone:(865) 946-1874 fax:(865) 946-1010 email: lrispoli at utk.edu
ADD COMMENTlink modified 10.2 years ago by James W. MacDonald47k • written 10.2 years ago by Louisa A Rispoli/AS/EXP/UTIA160
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gravatar for James W. MacDonald
10.2 years ago by
United States
James W. MacDonald47k wrote:
Hi Louisa, Louisa A Rispoli/AS/EXP/UTIA wrote: > Hi all- > > I am fairly new to bioconductor and am very excited about the example > described in the affycoretools vignette since 1) it would make my mentor > very happy (almost the exact situation that we have in the data set), 2) > seemed like a simple solution to a issue I was not sure how to approach and > 3) package seemed straightforward and easy to use. This is probably a > stupid question arising from the fact that I have yet to fully understand R > and bioconductor but I have tried to spend a better portion of the day > trying to figure where I am going wrong and having no luck. I am trying to > compare samples (two treatment groups) amplified two different ways and > see which differentially expressed genes are shared between the two > different methodologies. According to the PCA plot I need to compute the > expression values for each methodology separately and then combine the > data. I can not seem to create the new expression set properly to perform > the limma analysis. I have attached a text file with my R code. > > If anyone could point me in the right direction (either the mistake I am > makeing or even the vignette that I should read closer) I would appreciate > the help. Sorry if this seems very silly but we do not have anyone here > familiar with Bioconductor to go to for help. > > Thanks > > Louisa > > Sorry required a repost, forgot that attachments were stripped. > > R version 2.7.0 (2008-04-22) > Copyright (C) 2008 The R Foundation for Statistical Computing > ISBN 3-900051-07-0 >> library(affycoretools) > Loading required package: affy > Loading required package: Biobase > Loading required package: tools > > Welcome to Bioconductor > > Vignettes contain introductory material. To view, type > 'openVignette()'. To cite Bioconductor, see > 'citation("Biobase")' and for packages 'citation(pkgname)'. > > Loading required package: affyio > Loading required package: preprocessCore > Loading required package: limma > Loading required package: GOstats > Loading required package: graph > Loading required package: GO.db > Loading required package: AnnotationDbi > Loading required package: DBI > Loading required package: RSQLite > Loading required package: annotate > Loading required package: xtable > Loading required package: RBGL > Loading required package: Category > Loading required package: genefilter > Loading required package: survival > Loading required package: splines > Loading required package: biomaRt > Loading required package: RCurl > > Attaching package: 'biomaRt' > > > The following object(s) are masked from package:annotate : > > getGO > > Loading required package: gcrma > Loading required package: matchprobes > Loading required package: annaffy > Loading required package: KEGG.db > > Attaching package: 'annaffy' > > > The following object(s) are masked from package:RCurl : > > getURL > > Warning messages: > 1: package 'DBI' was built under R version 2.7.1 > 2: package 'RSQLite' was built under R version 2.7.1 > 3: package 'xtable' was built under R version 2.7.1 > >> pd <-read.AnnotatedDataFrame("pData.txt", sep="\t", header=TRUE, > row.names=1) >> dat <- ReadAffy(phenoData= pd) >> pData(dat) > amp trt > PolyC-1.CEL PolyA Ctrl > PolyC-2.CEL PolyA Ctrl > PolyC-3.CEL PolyA Ctrl > PolyC-4.CEL PolyA Ctrl > PolyC-5.CEL PolyA Ctrl > PolyC-6.CEL PolyA Ctrl > PolyC-7.CEL PolyA Ctrl > PolyC-8.CEL PolyA Ctrl > PolyHS-1.CEL PolyA HS > PolyHS-2.CEL PolyA HS > PolyHS-3.CEL PolyA HS > PolyHS-4.CEL PolyA HS > PolyHS-5.CEL PolyA HS > PolyHS-6.CEL PolyA HS > PolyHS-7.CEL PolyA HS > PolyHS-8.CEL PolyA HS > WTC-1.CEL WT Ctrl > WTC-2.CEL WT Ctrl > WTC-3.CEL WT Ctrl > WTC-4.CEL WT Ctrl > WTC-5.CEL WT Ctrl > WTC-6.CEL WT Ctrl > WTC-7.CEL WT Ctrl > WTC-8.CEL WT Ctrl > WTHS-1.CEL WT HS > WTHS-2.CEL WT HS > WTHS-3.CEL WT HS > WTHS-4.CEL WT HS > WTHS-5.CEL WT HS > WTHS-6.CEL WT HS > WTHS-7.CEL WT HS > WTHS-8.CEL WT HS > >> eset <-affystart(groups=rep(1:4, each = 8), groupsnames = > unique(paste(pData(pd)[,1],pData(pd)[,2], sep="-")), phenoData = pd) > Background correcting > Normalizing > Calculating Expression > >> eset1 <-affystart(filenames=list.celfiles()[1:16], plot = FALSE, > pca=FALSE) > Background correcting > Normalizing > Calculating Expression > >> eset2 <-affystart(filenames=list.celfiles()[17:32], plot=FALSE, > pca=FALSE) > Background correcting > Normalizing > Calculating Expression > >> eset <- new("ExpressionSet", exprs=cbind(exprs(eset1), exprs(eset2)), > phenoData = pd, annotation=annotation(eset1)) > Error in validObject(.Object) : > invalid class "ExpressionSet" object: sampleNames differ between > assayData and phenoData That's odd. What do you get for sampleNames(pd)? > >> eset1 > ExpressionSet (storageMode: lockedEnvironment) > assayData: 24128 features, 16 samples > element names: exprs > phenoData > sampleNames: PolyC-1.CEL, PolyC-2.CEL, ..., PolyHS-8.CEL (16 total) > varLabels and varMetadata description: > sample: arbitrary numbering > featureData > featureNames: AFFX-BioB-3_at, AFFX-BioB-5_at, ..., BtAffx.29968.1.S1_at > (24128 total) > fvarLabels and fvarMetadata description: none > experimentData: use 'experimentData(object)' > Annotation: bovine > >> eset2 > ExpressionSet (storageMode: lockedEnvironment) > assayData: 24128 features, 16 samples > element names: exprs > phenoData > sampleNames: WTC-1.CEL, WTC-2.CEL, ..., WTHS-8.CEL (16 total) > varLabels and varMetadata description: > sample: arbitrary numbering > featureData > featureNames: AFFX-BioB-3_at, AFFX-BioB-5_at, ..., BtAffx.29968.1.S1_at > (24128 total) > fvarLabels and fvarMetadata description: none > experimentData: use 'experimentData(object)' > Annotation: bovine > > "If we knew what we were doing, it wouldn't be called Research." - Albert > Einstein > > Louisa Rispoli, Ph.D. Reproductive Physiology > Department of Animal Science > University of Tennessee, Knoxville > A105 Johnson Animal Research and Teaching Unit > 1750 Alcoa Highway > Knoxville, TN 37920 > phone:(865) 946-1874 > fax:(865) 946-1010 > email: lrispoli at utk.edu > > _______________________________________________ > Bioconductor mailing list > Bioconductor at stat.math.ethz.ch > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor -- James W. MacDonald, M.S. Biostatistician Affymetrix and cDNA Microarray Core University of Michigan Cancer Center 1500 E. Medical Center Drive 7410 CCGC Ann Arbor MI 48109 734-647-5623
ADD COMMENTlink written 10.2 years ago by James W. MacDonald47k
Hi- Thank you for your help. I checked that sampleNames(pd) and looks like I think it should, but since I am a newbie I will leave it to you to tell me. Re-ran with 2.7.1 because when opened R2.7.0 this AM affycoretools not loading biomaRt for some reason. I think it may have to do that I updated packages but I have no clue. So uninstalled 2.7.0 then installed the 2.7.1 because could not get the link for 2.7.0 to function for me. I do get some additional warnings and errors with the 2.7.1 version that I did not see yesterday. When I do plotPCA the legend formed correctly so I am not sure what went wrong with the affystart. I appreciate help and any recommendations. Sincerely Louisa R version 2.7.1 (2008-06-23) Copyright (C) 2008 The R Foundation for Statistical Computing ISBN 3-900051-07-0 > library(affycoretools) Loading required package: affy Loading required package: Biobase Loading required package: tools Welcome to Bioconductor Vignettes contain introductory material. To view, type 'openVignette()'. To cite Bioconductor, see 'citation("Biobase")' and for packages 'citation(pkgname)'. Loading required package: affyio Loading required package: preprocessCore Loading required package: limma Loading required package: GOstats Loading required package: graph Loading required package: GO.db Loading required package: AnnotationDbi Loading required package: DBI Loading required package: RSQLite Loading required package: annotate Loading required package: xtable Loading required package: RBGL Loading required package: Category Loading required package: genefilter Loading required package: survival Loading required package: splines Loading required package: biomaRt Loading required package: RCurl Attaching package: 'biomaRt' The following object(s) are masked from package:annotate : getGO Loading required package: gcrma Loading required package: matchprobes Loading required package: annaffy Loading required package: KEGG.db Attaching package: 'annaffy' The following object(s) are masked from package:RCurl : getURL > pd <-read.AnnotatedDataFrame("pData.txt", sep="\t", header=TRUE, row.names=1) > dat <- ReadAffy(phenoData= pd) > pData(dat) amp trt PolyC-1.CEL PolyA Ctrl PolyC-2.CEL PolyA Ctrl PolyC-3.CEL PolyA Ctrl PolyC-4.CEL PolyA Ctrl PolyC-5.CEL PolyA Ctrl PolyC-6.CEL PolyA Ctrl PolyC-7.CEL PolyA Ctrl PolyC-8.CEL PolyA Ctrl PolyHS-1.CEL PolyA HS PolyHS-2.CEL PolyA HS PolyHS-3.CEL PolyA HS PolyHS-4.CEL PolyA HS PolyHS-5.CEL PolyA HS PolyHS-6.CEL PolyA HS PolyHS-7.CEL PolyA HS PolyHS-8.CEL PolyA HS WTC-1.CEL WT Ctrl WTC-2.CEL WT Ctrl WTC-3.CEL WT Ctrl WTC-4.CEL WT Ctrl WTC-5.CEL WT Ctrl WTC-6.CEL WT Ctrl WTC-7.CEL WT Ctrl WTC-8.CEL WT Ctrl WTHS-1.CEL WT HS WTHS-2.CEL WT HS WTHS-3.CEL WT HS WTHS-4.CEL WT HS WTHS-5.CEL WT HS WTHS-6.CEL WT HS WTHS-7.CEL WT HS WTHS-8.CEL WT HS > eset <-affystart(groups=rep(1:4, each = 8), groupsnames = unique(paste(pData(pd)[,1],pData(pd)[,2], sep="-")), phenoData = pd) Background correcting Normalizing Calculating Expression Please give the x-coordinate for a legend. Please give the y-coordinate for a legend. Error in method(paste("Density plot", plottype, sep = "."), height = height, : unused argument(s) (groupsnames = c("PolyA-Ctrl", "PolyA-HS", "WT- Ctrl", "WT-HS"), phenoData = <s4 object="" of="" class="" "annotateddataframe"="">) In addition: Warning messages: 1: In pca.legend(pca, groupnames, pch, col, x.coord = x.coord, y.coord = y.coord, : NAs introduced by coercion 2: In pca.legend(pca, groupnames, pch, col, x.coord = x.coord, y.coord = y.coord, : NAs introduced by coercion > eset1 <-affystart(filenames=list.celfiles()[1:16], plot = FALSE, pca=FALSE) Background correcting Normalizing Calculating Expression > eset2 <-affystart(filenames=list.celfiles()[17:32], plot=FALSE, pca=FALSE) Background correcting Normalizing Calculating Expression > eset <- new("ExpressionSet", exprs=cbind(exprs(eset1), exprs(eset2)), phenoData = pd, annotation=annotation(eset1)) Error in validObject(.Object) : invalid class "ExpressionSet" object: sampleNames differ between assayData and phenoData > sampleNames(pd) [1] "PolyC-1.cel" "PolyC-2.cel" "PolyC-3.cel" "PolyC-4.cel" "PolyC-5.cel" "PolyC-6.cel" "PolyC-7.cel" "PolyC-8.cel" [9] "PolyHS-1.cel" "PolyHS-2.cel" "PolyHS-3.cel" "PolyHS-4.cel" "PolyHS-5.cel" "PolyHS-6.cel" "PolyHS-7.cel" "PolyHS-8.cel" [17] "WTC-1.cel" "WTC-2.cel" "WTC-3.cel" "WTC-4.cel" "WTC-5.cel" "WTC-6.cel" "WTC-7.cel" "WTC-8.cel" [25] "WTHS-1.cel" "WTHS-2.cel" "WTHS-3.cel" "WTHS-4.cel" "WTHS-5.cel" "WTHS-6.cel" "WTHS-7.cel" "WTHS-8.cel" > sampleNames(eset1) [1] "PolyC-1.CEL" "PolyC-2.CEL" "PolyC-3.CEL" "PolyC-4.CEL" "PolyC-5.CEL" "PolyC-6.CEL" "PolyC-7.CEL" "PolyC-8.CEL" [9] "PolyHS-1.CEL" "PolyHS-2.CEL" "PolyHS-3.CEL" "PolyHS-4.CEL" "PolyHS-5.CEL" "PolyHS-6.CEL" "PolyHS-7.CEL" "PolyHS-8.CEL" > sampleNames(eset2) [1] "WTC-1.CEL" "WTC-2.CEL" "WTC-3.CEL" "WTC-4.CEL" "WTC-5.CEL" "WTC-6.CEL" "WTC-7.CEL" "WTC-8.CEL" "WTHS-1.CEL" "WTHS-2.CEL" [11] "WTHS-3.CEL" "WTHS-4.CEL" "WTHS-5.CEL" "WTHS-6.CEL" "WTHS-7.CEL" "WTHS-8.CEL" > eset <- rma(dat) Background correcting Normalizing Calculating Expression > plotPCA(eset, groups = rep(1:4, each =8), groupnames = unique(paste(pData(pd)[,1],pData(pd)[,2], sep="-")))
ADD REPLYlink written 10.2 years ago by Louisa A Rispoli/AS/EXP/UTIA160
Hi Louisa, Louisa A Rispoli/AS/EXP/UTIA wrote: > Hi- > > Thank you for your help. I checked that sampleNames(pd) and looks like I > think it should, but since I am a newbie I will leave it to you to tell me. > Re-ran with 2.7.1 because when opened R2.7.0 this AM affycoretools not > loading biomaRt for some reason. I think it may have to do that I updated > packages but I have no clue. So uninstalled 2.7.0 then installed the 2.7.1 > because could not get the link for 2.7.0 to function for me. I do get some > additional warnings and errors with the 2.7.1 version that I did not see > yesterday. When I do plotPCA the legend formed correctly so I am not sure > what went wrong with the affystart. I appreciate help and any > recommendations. > > Sincerely > > Louisa > > > R version 2.7.1 (2008-06-23) > Copyright (C) 2008 The R Foundation for Statistical Computing > ISBN 3-900051-07-0 > >> library(affycoretools) > Loading required package: affy > Loading required package: Biobase > Loading required package: tools > > Welcome to Bioconductor > > Vignettes contain introductory material. To view, type > 'openVignette()'. To cite Bioconductor, see > 'citation("Biobase")' and for packages 'citation(pkgname)'. > > Loading required package: affyio > Loading required package: preprocessCore > Loading required package: limma > Loading required package: GOstats > Loading required package: graph > Loading required package: GO.db > Loading required package: AnnotationDbi > Loading required package: DBI > Loading required package: RSQLite > Loading required package: annotate > Loading required package: xtable > Loading required package: RBGL > Loading required package: Category > Loading required package: genefilter > Loading required package: survival > Loading required package: splines > Loading required package: biomaRt > Loading required package: RCurl > > Attaching package: 'biomaRt' > > > The following object(s) are masked from package:annotate : > > getGO > > Loading required package: gcrma > Loading required package: matchprobes > Loading required package: annaffy > Loading required package: KEGG.db > > Attaching package: 'annaffy' > > > The following object(s) are masked from package:RCurl : > > getURL >> pd <-read.AnnotatedDataFrame("pData.txt", sep="\t", header=TRUE, > row.names=1) >> dat <- ReadAffy(phenoData= pd) >> pData(dat) > amp trt > PolyC-1.CEL PolyA Ctrl > PolyC-2.CEL PolyA Ctrl > PolyC-3.CEL PolyA Ctrl > PolyC-4.CEL PolyA Ctrl > PolyC-5.CEL PolyA Ctrl > PolyC-6.CEL PolyA Ctrl > PolyC-7.CEL PolyA Ctrl > PolyC-8.CEL PolyA Ctrl > PolyHS-1.CEL PolyA HS > PolyHS-2.CEL PolyA HS > PolyHS-3.CEL PolyA HS > PolyHS-4.CEL PolyA HS > PolyHS-5.CEL PolyA HS > PolyHS-6.CEL PolyA HS > PolyHS-7.CEL PolyA HS > PolyHS-8.CEL PolyA HS > WTC-1.CEL WT Ctrl > WTC-2.CEL WT Ctrl > WTC-3.CEL WT Ctrl > WTC-4.CEL WT Ctrl > WTC-5.CEL WT Ctrl > WTC-6.CEL WT Ctrl > WTC-7.CEL WT Ctrl > WTC-8.CEL WT Ctrl > WTHS-1.CEL WT HS > WTHS-2.CEL WT HS > WTHS-3.CEL WT HS > WTHS-4.CEL WT HS > WTHS-5.CEL WT HS > WTHS-6.CEL WT HS > WTHS-7.CEL WT HS > WTHS-8.CEL WT HS > >> eset <-affystart(groups=rep(1:4, each = 8), groupsnames = > unique(paste(pData(pd)[,1],pData(pd)[,2], sep="-")), phenoData = pd) > Background correcting > Normalizing > Calculating Expression > Please give the x-coordinate for a legend. > Please give the y-coordinate for a legend. > Error in method(paste("Density plot", plottype, sep = "."), height = > height, : > unused argument(s) (groupsnames = c("PolyA-Ctrl", "PolyA-HS", "WT- Ctrl", > "WT-HS"), phenoData = <s4 object="" of="" class="" "annotateddataframe"="">) > In addition: Warning messages: > 1: In pca.legend(pca, groupnames, pch, col, x.coord = x.coord, y.coord = > y.coord, : > NAs introduced by coercion > 2: In pca.legend(pca, groupnames, pch, col, x.coord = x.coord, y.coord = > y.coord, : > NAs introduced by coercion > > >> eset1 <-affystart(filenames=list.celfiles()[1:16], plot = FALSE, > pca=FALSE) > Background correcting > Normalizing > Calculating Expression >> eset2 <-affystart(filenames=list.celfiles()[17:32], plot=FALSE, > pca=FALSE) > Background correcting > Normalizing > Calculating Expression >> eset <- new("ExpressionSet", exprs=cbind(exprs(eset1), exprs(eset2)), > phenoData = pd, annotation=annotation(eset1)) > Error in validObject(.Object) : > invalid class "ExpressionSet" object: sampleNames differ between > assayData and phenoData >> sampleNames(pd) > [1] "PolyC-1.cel" "PolyC-2.cel" "PolyC-3.cel" "PolyC-4.cel" > "PolyC-5.cel" "PolyC-6.cel" "PolyC-7.cel" "PolyC-8.cel" > [9] "PolyHS-1.cel" "PolyHS-2.cel" "PolyHS-3.cel" "PolyHS-4.cel" > "PolyHS-5.cel" "PolyHS-6.cel" "PolyHS-7.cel" "PolyHS-8.cel" > [17] "WTC-1.cel" "WTC-2.cel" "WTC-3.cel" "WTC-4.cel" > "WTC-5.cel" "WTC-6.cel" "WTC-7.cel" "WTC-8.cel" > [25] "WTHS-1.cel" "WTHS-2.cel" "WTHS-3.cel" "WTHS-4.cel" > "WTHS-5.cel" "WTHS-6.cel" "WTHS-7.cel" "WTHS-8.cel" This is the problem. If you look at the sample names above, you can see that the extension is .cel, whereas below it is .CEL. Since R is case-sensitive, these names are all different. You need to change the sample names in the pd object to have capitalized CEL extensions. Best, Jim >> sampleNames(eset1) > [1] "PolyC-1.CEL" "PolyC-2.CEL" "PolyC-3.CEL" "PolyC-4.CEL" > "PolyC-5.CEL" "PolyC-6.CEL" "PolyC-7.CEL" "PolyC-8.CEL" > [9] "PolyHS-1.CEL" "PolyHS-2.CEL" "PolyHS-3.CEL" "PolyHS-4.CEL" > "PolyHS-5.CEL" "PolyHS-6.CEL" "PolyHS-7.CEL" "PolyHS-8.CEL" >> sampleNames(eset2) > [1] "WTC-1.CEL" "WTC-2.CEL" "WTC-3.CEL" "WTC-4.CEL" "WTC-5.CEL" > "WTC-6.CEL" "WTC-7.CEL" "WTC-8.CEL" "WTHS-1.CEL" "WTHS-2.CEL" > [11] "WTHS-3.CEL" "WTHS-4.CEL" "WTHS-5.CEL" "WTHS-6.CEL" "WTHS-7.CEL" > "WTHS-8.CEL" > > >> eset <- rma(dat) > Background correcting > Normalizing > Calculating Expression >> plotPCA(eset, groups = rep(1:4, each =8), groupnames = > unique(paste(pData(pd)[,1],pData(pd)[,2], sep="-"))) > -- James W. MacDonald, M.S. Biostatistician Affymetrix and cDNA Microarray Core University of Michigan Cancer Center 1500 E. Medical Center Drive 7410 CCGC Ann Arbor MI 48109 734-647-5623
ADD REPLYlink written 10.2 years ago by James W. MacDonald47k
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