In directory reports/CLP/ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6/cbs/
you should see many PNG files. Do they exists? How many are they? What do you see when you open them "manually"? What internet browser do you use? /Henrik On Mon, Jun 29, 2009 at 12:50 AM, Myriam Peyrard<[email protected]> wrote: > Thank you Henrik. > I re-run the whole until fragment normalization in the script I sent you > before (see below). > Then did: > plotDensity(cesN) > and seeing a nice number of curves, more or less overlapping for my arrays. > I attach the output (in 2 different file formats) and may be you can judge > yourself that it is as it should. > > So I have signals as I see density curves. > I run the analysis on my laptop which allows me to use 1.5 Gb Ram. I have > ordered one more Gb of memory but will have to wait a few days to get it > installed. If this is the problem, then I will have to wait end of July to > try it as I will be gone for 3 weeks from next week. > > But as I see data in my Excel regions files, isn´t so that the data is > produced and just the form of the output is the problem? > > Best regards, > Myriam. > > -----Original Message----- > From: [email protected] > [mailto:[email protected]] On Behalf Of Henrik Bengtsson > Sent: den 29 juni 2009 01:12 > To: [email protected] > Subject: [aroma.affymetrix] Re: No plot CNV analysis (Myriam) > > > Hi, > > my initial guess is that the when the fragmentation-length > normalization failed the first time it somehow generated empty output > files. To check this, try to plot the FLN normalized signals, e.g. > > plotDensity(cesN) > > You should see density curves as in > > > http://groups.google.com/group/aroma-affymetrix/web/estimation-of-total-copy > -numbers-using-the-crma-v2-method > > If not, the signals are probably all zeros. If so, then delete the > following directory: > > plmData/CLP,ACC,ra,-XY,AVG,+300,A+B,FLN,-XY/GenomeWideSNP_6 > > and retry your script. > > Also, you shouldn't run out of memory here unless you have very > limited RAM. Are you loading a large session image at the start? > > Cheers, > > /Henrik > > On Mon, Jun 22, 2009 at 1:07 AM, Myriam Peyrard<[email protected]> wrote: >> >> 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 >>>> >>>> >>>> >>>> >>>> >>>> >>>> > >>> >>> >>> >>> >>> >>> > >>> >> >> >> >> >> >> > >> > > > > > > --~--~---------~--~----~------------~-------~--~----~ When reporting problems on aroma.affymetrix, make sure 1) to run the latest version of the package, 2) to report the output of sessionInfo() and traceback(), and 3) to post a complete code example. 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