[BioC] RMA XPS Problem on MoGene 1.0 ST - Solved

[cstrato]

Dear Zack,

The probeset annotation file for the exon arrays contains a column 
"level", which was set to "" for whole genome arrays. In version na31 
Affymetrix has now set "level" to "---". This change caused the problem 
which is now solved in the new version xps_1.10.1 which is available 
from BioC 2.7.

Best regards
Christian


On 10/12/10 10:30 PM, cstrato wrote:
> Dear Zack,
>
> Meanwhile I could repeat your results when using the new annotation
> files version na31. However, when I use the old annotation files na30
> then everything is ok. This means that there may be a problem with the
> new annotation files. I need to investigate further and will let you
> know. Meanwhile I suggest that you use the annotation files na30.
>
> Best regards
> Christian
>
>
> On 10/11/10 9:55 PM, cstrato wrote:
>> Dear Zack,
>>
>> Usually you get this result when you create the scheme file for MoGene
>> using "import.genome.scheme" instead of using "import.exon.scheme", see
>> e.g.:
>> https://www.stat.math.ethz.ch/pipermail/bioconductor/2010-March/032353.html
>>
>>
>> However, it seems that you have created the scheme correctly. Thus could
>> you please send me the output of:
>> > str(scheme.moge10stv1r4.na31)
>>
>> Which version of the Affymetrix annotation files did you use to create
>> "scheme.moge10stv1r4.na31"?
>> You need to use the annotation files created on 09/08/10 and not the
>> ones created on 08/30/10 since the older files have additional
>> "control->affx" which are "neg_control".
>>
>> Another problem could be your code:
>> > data.rma <-rma(data.moge, ...., xps.scheme=scheme.moge10stv1r4.na31)
>>
>> Since "xps.scheme" should only be used if you want to use alternative
>> CDF-files for expression arrays, you should do:
>> > data.rma <-rma(data.moge, ...., xps.scheme=NULL)
>>
>> Please send me:
>> - your sessionInfo(),
>> - the output of str(scheme.moge10stv1r4.na31),
>> - the output of str(data.rma),
>> and let me know
>> - which annotation files you have used and
>> - whether rma(.., xps.scheme=NULL) solves the problem.
>>
>> Best regards
>> Christian
>> _._._._._._._._._._._._._._._._._._
>> C.h.r.i.s.t.i.a.n S.t.r.a.t.o.w.a
>> V.i.e.n.n.a A.u.s.t.r.i.a
>> e.m.a.i.l: cstrato at aon.at
>> _._._._._._._._._._._._._._._._._._
>>
>>
>> On 10/11/10 5:07 PM, Zack Liu wrote:
>>> Dear members,
>>>
>>> I have encountered some problems using XPS library on Mouse Gene 1.0 ST
>>> arrays.. Basically, when I run rma on my cel files, the signal matrix
>>> only
>>> have 21 rows (probeset?) for all the samples. Has anybody here had the
>>> same
>>> problem before?
>>>
>>> ## Generate the Scheme file
>>>
>>> scmdir<- paste(.path.package("xps"),"schemes",sep="/")
>>>
>>> libdir<- "./Affy/libraryfiles"
>>>
>>> anndir<- "./Affy/Annotation"
>>>
>>>
>>> scheme.moge10stv1r4.na31<-
>>> import.exon.scheme("Scheme_MoGe10stv1r4_na31",
>>> filedir=scmdir, layoutfile=paste(libdir,
>>> "MoGene-1_0-st-v1.r4.clf",sep="/"),schemefile=paste(libdir,
>>> "MoGene-1_0-st-v1.r4.pgf",sep="/"), probeset=paste(anndir,
>>> "MoGene-1_0-st-v1.na31.mm9.probeset.csv",sep="/"),
>>> transcript=paste(anndir,"MoGene-1_0-st-v1.na31.mm9.transcript.csv",sep="/"))
>>>
>>>
>>>
>>>
>>> ### Generate the signal file
>>>
>>>
>>> celDir<- "./rawData/tissues/MoGene-1-0-st-v1/"
>>>
>>> celfiles<- c("001.CEL","002.CEL","003.CEL")
>>>
>>> celNames<- c("01","02","03")
>>>
>>> datdir<-"rootData"
>>>
>>>
>>> data.moge<- import.data(scheme.moge10stv1r4.na31,"mogene_glio",filedir=
>>> datdir,celdir=celDir,celfiles=celfiles,celnames =celNames,verbose=T)
>>>
>>>
>>> data.moge<- attachInten(data.moge)
>>>
>>> tmp<- intensity(data.moge)
>>>
>>> head(tmp)
>>>
>>>> dim(tmp)
>>>
>>> [1] 1102500 5
>>>
>>>
>>> ### So far so good, I have
>>>
>>> #### Problem starts here
>>>
>>> data.rma<-rma(data.moge, "tmp_MoGene_Glio_RMA",filedir=datdir,tmpdir="",
>>> option ="transcript", background="antigenomic",normalize=TRUE,exonlevel=
>>> "all",verbose=TRUE,xps.scheme=scheme.moge10stv1r4.na31)
>>>
>>>
>>> Creating new temporary file<rootData/tmp_MoGene_Glio_RMA.root>...
>>>
>>> Opening file
>>> </Library/Frameworks/R.framework/Resources/library/xps/schemes/Scheme_MoGe10stv1r4_na31.root>
>>>
>>>
>>> in<READ> mode...
>>>
>>> Opening file<./rootData/mogene_glio_cel.root> in<READ> mode...
>>>
>>> Added<3> trees to PreprocesSet.
>>>
>>> Preprocessing data using method<preprocess>...
>>>
>>> Background correcting raw data...
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> calculating background for<01.cel>...
>>>
>>> background statistics:
>>>
>>> 1087986 cells with minimal intensity 0
>>>
>>> 970 cells with maximal intensity 84.8093
>>>
>>> calculating background for<02.cel>...
>>>
>>> background statistics:
>>>
>>> 1087986 cells with minimal intensity 0
>>>
>>> 4596 cells with maximal intensity 82.8614
>>>
>>> calculating background for<03.cel>...
>>>
>>> background statistics:
>>>
>>> 1087986 cells with minimal intensity 0
>>>
>>> 3018 cells with maximal intensity 71.3567
>>>
>>> Normalizing raw data...
>>>
>>> normalizing data using method<quantile>...
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> finished filling<3> arrays.
>>>
>>> computing common mean...
>>>
>>> finished filling<3> trees.
>>>
>>> Converting raw data to expression levels...
>>>
>>> summarizing with<medianpolish>...
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> calculating expression for<21> of<35556> units...Finished.
>>>
>>> expression statistics:
>>>
>>> minimal expression level is<36.657>
>>>
>>> maximal expression level is<14222.3>
>>>
>>> preprocessing finished.
>>>
>>> Opening file
>>> </Library/Frameworks/R.framework/Resources/library/xps/schemes/Scheme_MoGe10stv1r4_na31.root>
>>>
>>>
>>> in<READ> mode...
>>>
>>> Opening file<rootData/tmp_MoGene_Glio_RMA.root> in<READ> mode...
>>>
>>> Opening file<rootData/tmp_MoGene_Glio_RMA.root> in<READ> mode...
>>>
>>> Exporting data from tree<*> to file<rootData/tmp_MoGene_Glio_RMA.txt>...
>>>
>>> Reading entries from<MoGene-1_0-st-v1.ann> ...Finished
>>>
>>> <21> of<21> records exported.
>>>
>>>>
>>>
>>>>
>>>
>>>>
>>>
>>>> dim(tmp)
>>>
>>> Error: object 'tmp' not found
>>>
>>>>
>>>
>>>> data.moge<- attachInten(data.moge)
>>>
>>> tmp<- intensity(data.moge)
>>>
>>> head(tmp)
>>>
>>>
>>>
>>>
>>>> tmp<- intensity(data.moge)
>>>
>>>> head(tmp)
>>>
>>> X Y 01.cel_MEAN 02.cel_MEAN 03.cel_MEAN
>>>
>>> 1 0 0 6213 8575 6339
>>>
>>> 2 1 0 105 198 126
>>>
>>> 3 2 0 6361 8720 6278
>>>
>>> 4 3 0 138 170 120
>>>
>>> 5 4 0 138 180 134
>>>
>>> 6 5 0 127 139 133
>>>
>>>>
>>>
>>>>
>>>
>>>>
>>>
>>>> dim(tmp)
>>>
>>> [1] 1102500 5
>>>
>>>>
>>>
>>>>
>>>
>>>> data.rma<-rma(data.moge,
>>> "tmp_MoGene_Glio_RMA",filedir=datdir,tmpdir="",option ="transcript",
>>> background="antigenomic",normalize=TRUE,exonlevel="all",verbose=TRUE,xps.scheme=scheme.moge10stv1r4.na31)
>>>
>>>
>>>
>>> Creating new temporary file<rootData/tmp_MoGene_Glio_RMA.root>...
>>>
>>> Opening file
>>> </Library/Frameworks/R.framework/Resources/library/xps/schemes/Scheme_MoGe10stv1r4_na31.root>
>>>
>>>
>>> in<READ> mode...
>>>
>>> Opening file<rootData/mogene_glio_cel.root> in<READ> mode...
>>>
>>> Added<3> trees to PreprocesSet.
>>>
>>> Preprocessing data using method<preprocess>...
>>>
>>> Background correcting raw data...
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> calculating background for<01.cel>...
>>>
>>> background statistics:
>>>
>>> 1087986 cells with minimal intensity 0
>>>
>>> 970 cells with maximal intensity 84.8093
>>>
>>> calculating background for<02.cel>...
>>>
>>> background statistics:
>>>
>>> 1087986 cells with minimal intensity 0
>>>
>>> 4596 cells with maximal intensity 82.8614
>>>
>>> calculating background for<03.cel>...
>>>
>>> background statistics:
>>>
>>> 1087986 cells with minimal intensity 0
>>>
>>> 3018 cells with maximal intensity 71.3567
>>>
>>> Normalizing raw data...
>>>
>>> normalizing data using method<quantile>...
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> finished filling<3> arrays.
>>>
>>> computing common mean...
>>>
>>> finished filling<3> trees.
>>>
>>> Converting raw data to expression levels...
>>>
>>> summarizing with<medianpolish>...
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> setting selector mask for typepm<16316>
>>>
>>> calculating expression for<21> of<35556> units...Finished.
>>>
>>> expression statistics:
>>>
>>> minimal expression level is<36.657>
>>>
>>> maximal expression level is<14222.3>
>>>
>>> preprocessing finished.
>>>
>>> Opening file
>>> </Library/Frameworks/R.framework/Resources/library/xps/schemes/Scheme_MoGe10stv1r4_na31.root>
>>>
>>>
>>> in<READ> mode...
>>>
>>> Opening file<rootData/tmp_MoGene_Glio_RMA.root> in<READ> mode...
>>>
>>> Opening file<rootData/tmp_MoGene_Glio_RMA.root> in<READ> mode...
>>>
>>> Exporting data from tree<*> to file<rootData/tmp_MoGene_Glio_RMA.txt>...
>>>
>>> Reading entries from<MoGene-1_0-st-v1.ann> ...Finished
>>>
>>> <21> of<21> records exported.
>>>
>>>
>>>> expr.rma<- validData(data.rma)
>>>
>>>> dim(expr.rma)
>>>
>>> [1] 21 3
>>>
>>>
>>> ## only 21 records....
>>>
>>>
>>> Can anyone help? Thank you!
>>>
>>>
>>> Zach Liu
>>>
>>>
>>> Genomics& Computational Biology Program
>>> Abramson Cancer Institute, School of Medicine
>>> University of Pennsylvania
>>>
>>> [[alternative HTML version deleted]]
>>>
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>>
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