Dear Henrik,
I do not get any result, on any chromosomes. But I get all the data in Excel
format in the regions.xls file. Just that it is hard to compile without the
visualisation. I am using the vignette Total Copy Number Analysis (5.0 &
6.0). Myriam
Here is my session, from my notes in Tinn-R:
library(aroma.affymetrix)
library(Biobase)
library(tools)
chipType <- "GenomeWideSNP_6"
verbose <- Arguments$getVerbose(-8)
timestampOn(verbose)
cdf <- AffymetrixCdfFile$byChipType("GenomeWideSNP_6", tags="Full")
cdf
gi <- getGenomeInformation(cdf)
gi
si <- getSnpInformation(cdf)
#######################################################################
#Defining the CEL-set
#######################################################################
cs <- AffymetrixCelSet$byName("CLP", cdf=cdf)
cs
#AffymetrixCelSet:
#Name: CLP
#Tags:
#Path: rawData/CLP/GenomeWideSNP_6
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full
#Number of arrays: 28
#Names: MP01_LKG12, MP02_LKG15, ..., MP28_LKG226
#Time period: 2009-02-04 21:40:20 -- 2009-03-11 14:10:37
#Total file size: 1844.47MB
#RAM: 0.02MB
#######################################################################
#Calibration for allelic crosstalk
#######################################################################
acc <- AllelicCrosstalkCalibration(cs)
acc
#AllelicCrosstalkCalibration:
#Data set: CLP
#Input tags:
#User tags: *
#Asterisk ('*') tags: ACC,ra,-XY
#Output tags: ACC,ra,-XY
#Number of files: 28 (1844.47MB)
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full
#Algorithm parameters: (rescaleBy: chr "all", targetAvg: num 2200,
subsetToAvg: chr "-XY", mergeShifts: logi TRUE, B: int 1, flavor: chr
"sfit", algorithmParameters:List of 3, ..$ alpha: num [1:5] 0.1 0.075 0.05
0.03 0.01, ..$ q: num 2, ..$ Q: num 98)
#Output path: probeData/CLP,ACC,ra,-XY/GenomeWideSNP_6
#Is done: FALSE
#RAM: 0.00MB
csC <- process(acc, verbose=verbose)
csC
#AffymetrixCelSet:
#Name: CLP
#Tags: ACC,ra,-XY
#Path: probeData/CLP,ACC,ra,-XY/GenomeWideSNP_6
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full
#Number of arrays: 28
#Names: MP01_LKG12, MP02_LKG15, ..., MP28_LKG226
#Time period: 2009-06-08 11:01:00 -- 2009-06-08 12:51:15
#Total file size: 1840.64MB
#RAM: 0.02MB
#######################################################################
#Summarization
#######################################################################
plm <- AvgCnPlm(csC, mergeStrands=TRUE, combineAlleles=TRUE, shift=+300)
plm
#AvgCnPlm:
#Data set: CLP
#Chip type: GenomeWideSNP_6,Full
#Input tags: ACC,ra,-XY
#Output tags: ACC,ra,-XY,AVG,+300,A+B
#Parameters: (probeModel: chr "pm"; shift: num 300; flavor: chr "median";
mergeStrands: logi TRUE; combineAlleles: logi TRUE).
#Path: plmData/CLP,ACC,ra,-XY,AVG,+300,A+B/GenomeWideSNP_6
#RAM: 0.00MB
if (length(findUnitsTodo(plm)) > 0) {
# Fit CN probes quickly (~5-10s/array + some overhead)
units <- fitCnProbes(plm, verbose=verbose)
str(units)
# int [1:945826] 935590 935591 935592 935593 935594 935595 ...
# Fit remaining units, i.e. SNPs (~5-10min/array)
units <- fit(plm, verbose=verbose)
str(units)
}
ces <- getChipEffectSet(plm)
ces
#CnChipEffectSet:
#Name: CLP
#Tags: ACC,ra,-XY,AVG,+300,A+B
#Path: plmData/CLP,ACC,ra,-XY,AVG,+300,A+B/GenomeWideSNP_6
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full,monocell
#Number of arrays: 28
#Names: MP01_LKG12, MP02_LKG15, ..., MP28_LKG226
#Time period: 2009-06-08 15:13:31 -- 2009-06-08 15:13:35
#Total file size: 754.58MB
#RAM: 0.03MB
#Parameters: (probeModel: chr "pm", mergeStrands: logi TRUE, combineAlleles:
logi TRUE)
#######################################################################
#PCR fragment length normalization
#######################################################################
fln <- FragmentLengthNormalization(ces)
fln
#FragmentLengthNormalization:
#Data set: CLP
#Input tags: ACC,ra,-XY,AVG,+300,A+B
#User tags: *
#Asterisk ('*') tags: FLN,-XY
#Output tags: ACC,ra,-XY,AVG,+300,A+B,FLN,-XY
#Number of files: 28 (754.58MB)
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full,monocell
#Algorithm parameters: (subsetToFit: chr "-XY", onMissing: chr "median",
.target: NULL, shift: num 0)
#Output path: plmData/CLP,ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6
#Is done: FALSE
#RAM: 0.00MB
cesN <- process(fln, verbose=verbose)
#There were 29 warnings (use warnings() to see them)
#> warnings()
#Error: cannot allocate vector of size 14.3 Mb
#In addition: Warning messages:
#1: In attributes(.Data) <- c(attributes(.Data), attrib) :
# Reached total allocation of 1535Mb: see help(memory.size)
#2: In attributes(.Data) <- c(attributes(.Data), attrib) :
# Reached total allocation of 1535Mb: see help(memory.size)
#3: In attributes(.Data) <- c(attributes(.Data), attrib) :
# Reached total allocation of 1535Mb: see help(memory.size)
#4: In attributes(.Data) <- c(attributes(.Data), attrib) :
# Reached total allocation of 1535Mb: see help(memory.size)
#checking memory allocation, limit of computer and incresing to the max one
can use now.
memory.size()
#[1] 1283.22
memory.limit(size = NA)
#[1] 2000
memory.size(max = TRUE)
#[1] 1527.44
#RE-DID STEP BEFORE WHERE 29 WARNINGS AND PROBLEM WITH MEMORY
cesN <- process(fln, verbose=verbose)
#20090609 15:23:58|Normalizing set for PCR fragment-length effects...
#20090609 15:23:59| Already normalized
#20090609 15:23:59| Getting output data set...
#20090609 15:23:59| Getting output data set for
FragmentLengthNormalization...
#20090609 15:23:59| Calling NextMethod:
# List of 6
# $ generic : chr "getOutputDataSet"
# $ :Classes 'FragmentLengthNormalization',
'ChipEffectTransform', 'Transform', 'AromaTransform', 'Object' atomic [1:1]
NA
# .. ..- attr(*, ".env")=<environment: 0x0ae9f108>
# .. ..- attr(*, "...instantiationTime")= POSIXct[1:1], format:
"2009-06-09 09:34:09"
# $ cdf :Classes 'AffymetrixCdfFile', 'UnitNamesFile',
'AromaChipTypeAnnotationFile', 'AffymetrixFile', 'AromaPlatformInterface',
'AromaMicroarrayDataFile', 'GenericDataFile', 'Object' atomic [1:1] NA
# .. ..- attr(*, ".env")=<environment: 0x0a0bcc6c>
# .. ..- attr(*, "...instantiationTime")= POSIXct[1:1], format:
"2009-06-08 15:13:31"
# $ checkChipType : logi FALSE
# $ combineAlleles: logi TRUE
# $ mergeStrands : logi TRUE
#20090609 15:23:59| Getting output data set for
FragmentLengthNormalization...
#20090609 15:23:59| Already retrived.
#20090609 15:23:59| Getting output data set for
FragmentLengthNormalization...done
#20090609 15:23:59| Updating CnChipEffectSet...
#20090609 15:23:59| Scanning for and applying sample annotation files...
#20090609 15:23:59| Defining 0 files...
#20090609 15:23:59| Defining 0 files...done
#20090609 15:23:59| No sample annotation files found.
#20090609 15:23:59| Scanning for and applying sample annotation
files...done
#20090609 15:23:59| Updating CnChipEffectSet...done
#20090609 15:23:59| Getting output data set for
FragmentLengthNormalization...done
#20090609 15:23:59| Getting output data set...done
#20090609 15:23:59|Normalizing set for PCR fragment-length effects...done
cesN
#CnChipEffectSet:
#Name: CLP
#Tags: ACC,ra,-XY,AVG,+300,A+B,FLN,-XY
#Path: plmData/CLP,ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full,monocell
#Number of arrays: 28
#Names: MP01_LKG12, MP02_LKG15, ..., MP28_LKG226
#Time period: 2009-06-08 15:13:31 -- 2009-06-08 15:13:35
#Total file size: 754.58MB
#RAM: 0.03MB
#Parameters: (probeModel: chr "pm", mergeStrands: logi TRUE, combineAlleles:
logi TRUE)
#######################################################################
#Identification of copy-number regions (CNRs)
#######################################################################
cbs <- CbsModel(cesN)
#Loading required package: DNAcopy
cbs
#CbsModel:
#Name: CLP
#Tags: ACC,ra,-XY,AVG,+300,A+B,FLN,-XY
#Chip type (virtual): GenomeWideSNP_6
#Path: cbsData/CLP,ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6
#Number of chip types: 1
#Chip-effect set & reference file pairs:
#Chip type #1 of 1 ('GenomeWideSNP_6'):
#Chip-effect set:
#CnChipEffectSet:
#Name: CLP
#Tags: ACC,ra,-XY,AVG,+300,A+B,FLN,-XY
#Path: plmData/CLP,ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6
#Platform: Affymetrix
#Chip type: GenomeWideSNP_6,Full,monocell
#Number of arrays: 28
#Names: MP01_LKG12, MP02_LKG15, ..., MP28_LKG226
#Time period: 2009-06-08 15:13:31 -- 2009-06-08 15:13:35
#Total file size: 754.58MB
#RAM: 0.03MB
#Parameters: (probeModel: chr "pm", mergeStrands: logi TRUE, combineAlleles:
logi TRUE)
#Reference file:
#<average across arrays>
#RAM: 0.00MB
#######################################################################
#Fitting copy-number model and displaying results
#######################################################################
ce <- ChromosomeExplorer(cbs)
ce
#ChromosomeExplorer:
#Name: CLP
#Tags: ACC,ra,-XY,AVG,+300,A+B,FLN,-XY
#Number of arrays: 28
#Path: reports/CLP/ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6/cbs
#RAM: 0.00MB
process(ce,
chromosomes=c(1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23),
verbose=verbose)
display(ce)
-----Original Message-----
From: [email protected]
[mailto:[email protected]] On Behalf Of Henrik Bengtsson
Sent: den 18 juni 2009 18:25
To: [email protected]
Subject: Re: No CN estimates for SNP 6.0 (Was: Re: [aroma.affymetrix] Re:
CNV analys 28 st SNP 6.0 arrays)
Hi,
are you saying you do not get results from any chromosome? If so,
that's a first very useful clue; if you get from some, something odd
is going odd. Doing CN analysis on these chips is standard procedure,
so it "should work" out of the box.
In order to help you, what version are you using, i.e. output of
sessionInfo(). Exactly what is script you are using?
/Henrik
> -----Original Message-----
> From: Myriam Peyrard
> Sent: den 16 juni 2009 17:01
> To: 'aroma-affymetrix'
> Subject: RE: [aroma.affymetrix] Re: CNV analys 28 st SNP 6.0 arrays
>
> I have not gotten any answer about what can be wrong in my Aroma analysis
> when I cannot visualize any CNVs or else on my chromsome maps in
> ChromosomeExplorer. I tried now to run only one chromsome (nr 19) and
again,
> nothing on the plots even for 19.
> I then looked at the Excel file in Cbs named "regions" and there are data
in
> there. So the analysis seems to work but not the visualization.
> What shall I do/try? Is it a computer memory, capacity problem?
> ChromosomeExplorer compatibility?
>
> Myriam
On Thu, Jun 11, 2009 at 1:11 AM, Myriam Peyrard<[email protected]> wrote:
>
> Moreover, chrom 1 and 2, in all samples, seems not to have been done, (in
> grey shade when browsing acrross chromosomes and samples.
> Myriam
>
> -----Original Message-----
> From: Myriam Peyrard [mailto:[email protected]]
> Sent: den 11 juni 2009 10:09
> To: '[email protected]'
> Subject: RE: [aroma.affymetrix] Re: CNV analys 28 st SNP 6.0 arrays
> Importance: High
>
> OK, nice to hear, about the warnings.
> Then I do:
> display(ce)
> and started to look in my different samples, all chromosomes.
> Seems that there is not a single CNV variation there?
> I did the normalization as non-paired sample sets, with average calculated
> with all samples:
>
> cbs <- CbsModel(cesN)
>
> Is this a normal output? I expected to have many variations and to have to
> sort them out in different ways. Now, I think something has gone wrong
> instead. Any ideas?
>
> Myriam
>
> -----Original Message-----
> From: [email protected]
> [mailto:[email protected]] On Behalf Of Henrik Bengtsson
> Sent: den 11 juni 2009 09:38
> To: [email protected]
> Subject: [aroma.affymetrix] Re: CNV analys 28 st SNP 6.0 arrays
>
>
> Hi.
>
> On Thu, Jun 11, 2009 at 12:29 AM, Myriam <[email protected]> wrote:
>>
>> Dear all,
>>
>> I have gone through a number of steps in the analysis of 28 arrays of
>> the Genomewide6.0 type and all seems OK until now:
>>
>> #Fitting copy-number model and displaying results
>> #######################################################################
>> ce <- ChromosomeExplorer(cbs)
>> ce
>>
>> #ChromosomeExplorer:
>> #Name: CLP
>> #Tags: ACC,ra,-XY,AVG,+300,A+B,FLN,-XY
>> #Number of arrays: 28
>> #Path: reports/CLP/ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6/cbs
>> #RAM: 0.00MB
>>
>> process(ce, chromosomes=c
>> (1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,18,19,20,21,22,23),
>> verbose=verbose)
>
> FYI, you can write the above much shorter:
>
> process(ce, chromosomes=1:23, verbose=verbose)
>
>>
>> And the end of the output after many hours run, looks like this:
>> #20090610 02:17:03| Writing CN regions...done
>> #20090610 02:17:03|Generating ChromosomeExplorer report...done
>> #[1] TRUE
>> #There were 50 or more warnings (use warnings() to see the first 50)
>>
>> I am looking at the 50 first warnings and I see that many are of the
>> same type. here comes an example of one of the each category of
>> warning. What shall I do now with each of those?
>
> You don't have to do anything. These warnings are alright and
> expected. No worries here. If you on the other hand get *errors*
> then you have to worry, but those will halt the script immediately.
>
> /Henrik
>
>> 1: In library(package, lib.loc = lib.loc, character.only =
>> TRUE, ... :
>> there is no package called 'Cairo'
>> 2: In method(static, ...) :
>> Ghostscript not found. Searched directories: C:/gs, C:\Program/gs, /
>> gs, C:\Program\Delade filer/gs
>> 3: In DNAcopy::CNA(genomdat = data$y, chrom = data$chromosome, ... :
>> array has repeated maploc positions
>>
>> 4: In plotDev(pathname, width = width, height = height) :
>> Unable to allocate bitmap
>> 5: In plotDev(pathname, width = width, height = height) : opening
>> device failed
>> 6: In log(theta/thetaRef, base = 2) : NaNs produced
>> 7: In log(theta * thetaRef, base = 2) : NaNs produced
>> 8: In DNAcopy::CNA(genomdat = data$y, chrom = data$chromosome, ... :
>> array has repeated maploc positions
>>
>> Thanks a lot in adavnce for any help!
>> Myriam
>>
>>
>>
>>
>>
>>
>> >
>
>
>
>
>
> >
>
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