Entering edit mode
@louisa-a-rispoliasexputia-2436
Last seen 9.7 years ago
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
