java.lang.IndexOutOfBoundsException & Exception in thread "Thread-25" java.lang.StackOverflowError

[pswaminath]

Hi, I am using JACUSA for my project. I have gDNA (from bwa aligner) and cDNA (from STAR aligner) bam files sorted and indexed. I used the following command with java 1.7. (hs.bed contains chromosome 1 to Y)

java -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1,D -b hs37d5.bed -p 26 -r rddsnov27aoption.out -s gDNA.bam cDNA.bam &> jacnov27aoption.log

then I got the following errors. The process was stuck and there was no output but only tmp.gz files

java.lang.IndexOutOfBoundsException: Index: 0, Size: 0 at java.util.ArrayList.rangeCheck(ArrayList.java:635) at java.util.ArrayList.get(ArrayList.java:411) at java.util.Collections$UnmodifiableList.get(Collections.java:1211) at jacusa.filter.storage.DistanceFilterStorage.processRecord(DistanceFilterStorage.java:39) at jacusa.pileup.builder.AbstractPileupBuilder.processRecord(AbstractPileupBuilder.java:358) at jacusa.pileup.builder.AbstractPileupBuilder.adjustWindowStart(AbstractPileupBuilder.java:178) at jacusa.pileup.iterator.AbstractWindowIterator.adjustWindowStart(AbstractWindowIterator.java:155) at jacusa.pileup.iterator.AbstractWindowIterator.adjustCurrentGenomicPosition(AbstractWindowIterator.java:148) at jacusa.pileup.iterator.TwoSampleIterator.hasNext(TwoSampleIterator.java:38) at jacusa.pileup.worker.AbstractWorker.processParallelPileupIterator(AbstractWorker.java:183) at jacusa.pileup.worker.AbstractWorker.run(AbstractWorker.java:67)

Exception in thread "Thread-25" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

Complete error log attached as pdf below.

jacusaerrorforgitissue.pdf

Please suggest as how these errors could be resolved. Thank you! Priya

[piechottam]

Hi,

according to your Exception [...] at jacusa.filter.storage.DistanceFilterStorage.processRecord(DistanceFilterStorage.java:39) [...] means that there are no alignment blocks for some read - this is strange. This basically means, there is a read marked as mapped but it does not have any alignment blocks.

"-b hs37d5.bed" You are specifically looking for reads mapping to the decoy genome (hs37d5)? Do gDNA.bam cDNA.bam contain all reads? Was the alignment against (human genome + hs37d5) or first against human genome and then unmapped against hs37d5? Do you get the error if you run JACUSA on human chromosomes 1, - 23, X, Y, MT? How big are your files? It is hard to help without looking at the data.

Best, Michael

Am Mi., 27. Nov. 2019 um 21:31 Uhr schrieb pswaminath < notifications@github.com>:

Hi, I am using JACUSA for my project. I have gDNA (from bwa aligner) and cDNA (from STAR aligner) bam files sorted and indexed. I used the following command with java 1.7. (hs.bed contains chromosome 1 to Y)

java -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1,D -b hs37d5.bed -p 26 -r rddsnov27aoption.out -s gDNA.bam cDNA.bam &> jacnov27aoption.log

then I got the following errors. The process was stuck and there was no output but only tmp.gz files

java.lang.IndexOutOfBoundsException: Index: 0, Size: 0 at java.util.ArrayList.rangeCheck(ArrayList.java:635) at java.util.ArrayList.get(ArrayList.java:411) at java.util.Collections$UnmodifiableList.get(Collections.java:1211) at jacusa.filter.storage.DistanceFilterStorage.processRecord(DistanceFilterStorage.java:39) at jacusa.pileup.builder.AbstractPileupBuilder.processRecord(AbstractPileupBuilder.java:358) at jacusa.pileup.builder.AbstractPileupBuilder.adjustWindowStart(AbstractPileupBuilder.java:178) at jacusa.pileup.iterator.AbstractWindowIterator.adjustWindowStart(AbstractWindowIterator.java:155) at jacusa.pileup.iterator.AbstractWindowIterator.adjustCurrentGenomicPosition(AbstractWindowIterator.java:148) at jacusa.pileup.iterator.TwoSampleIterator.hasNext(TwoSampleIterator.java:38) at jacusa.pileup.worker.AbstractWorker.processParallelPileupIterator(AbstractWorker.java:183) at jacusa.pileup.worker.AbstractWorker.run(AbstractWorker.java:67)

Exception in thread "Thread-25" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

Complete error log attached as pdf below.

jacusaerrorforgitissue.pdf https://github.com/dieterich-lab/JACUSA/files/3898673/jacusaerrorforgitissue.pdf

Please suggest as how these errors could be resolved. Thank you! Priya

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[pswaminath]

Hi Michael, Thank you for your response. Here are the answers:

according to your Exception [...] at jacusa.filter.storage.DistanceFilterStorage.processRecord(DistanceFilterStorage.java:39) [...] means that there are no alignment blocks for some read - this is strange. This basically means, there is a read marked as mapped but it does not have any alignment blocks.

Thanks for identifying this error.

"-b hs37d5.bed" You are specifically looking for reads mapping to the decoy genome (hs37d5)? Yes. hs37d5.bed contains positions in chromosomes 1 though 23, X and Y

Do gDNA.bam cDNA.bam contain all reads? Was the alignment against (human genome + hs37d5) or first against human genome and then unmapped against hs37d5?

bam files were got after aligning with hs37d5 (GRch37+decoy). hs37d5.fa. GRCh37 primary assembly, mitochondrial sequences, GL000... seqs, Human herpesvirus (NC_007605) and the decoy sequences

Do you get the error if you run JACUSA on human chromosomes 1, - 23, X, Y, MT? Yes I got error when I ran chromosomes 1 through 23, X, Y (alignments restricted to chromosomes 1 through 23, X, Y in bam file) with hs37d5.bed and the process got stuck the same way. I did not check including MT.

Other test run that I did: When I ran JACUSA confined to regions of interest (chromosome1 only, chromosome Y only) individually using -b option, run was completed.

How big are your files? Bam files range in size from 6 - 8 GB

other data details ( it is paired end 2x150bp length)

It is hard to help without looking at the data.

Hope this helps. Please let me know if you need any other information. I appreciate your help.

Priya

[pswaminath]

Hi Michael, More information about our data: DNA library prep: agilent clinical research exome V1, 2x125bp RNA library prep: Illumina RNA Access, 2x125 bp unstranded

Thanks! Priya

[piechottam]

Hi,

thanks for sharing your details. Did you manage to get it to work?

Best, Michael

Am Fr., 29. Nov. 2019 um 21:04 Uhr schrieb pswaminath < notifications@github.com>:

Hi Michael, More information about our data: DNA library prep: agilent clinical research exome V1, 2x125bp RNA library prep: Illumina RNA Access, 2x125 bp unstranded

Thanks! Priya

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[pswaminath]

Thanks Michael for checking! I have explored the options that I knew but I haven't got it to work. Any help will be appreciated.

Priya

[piechottam]

It is really difficult without having the data. Can u pinpoint reads that cause the problem? Or at least the contig?

Am Mo., 2. Dez. 2019 um 16:05 Uhr schrieb pswaminath < notifications@github.com>:

Thanks Michael for checking! I have explored the options that I knew but I haven't got it to work. Any help will be appreciated.

Priya

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[pswaminath]

Hi Michael,

I ran JACUSA with the following command (complete bed file with positions in chr 1 to Y, bam file contains alignments chr 1 to Y)

Command used:

java -jar JACUSA_v1.3.0.jar call-2 -b hs37d5.bed -p 26 -r rddsdec2chr1to22XYNOaoption.out -s gDNAchr1to22andXY.bam TRNASTARwithMDchr1to22andXY.bam &> jacdec2chr1to22XYnooption.log

I have the complete log file, screenshot of tmp.gz files and the last lines of bed file attached here.

jacdec2chr1to22XYNOaoption.log

Please take a look.

I noticed that JACUSA run goes up to the positions in chr Y in the bed file provided by us and then it is stuck before wrapping up the result. Please confirm. It did not stop anywhere inbetween.

I have copied few lines where the error started and then the end of the messages. Please see attached log and screenshots for more information

[ INFO ] 05:19:44 : Started screening contig Y:28680525-28780524 [ INFO ] 05:19:44 : Started screening contig Y:58999981-59034049 Exception in thread "Thread-21" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

Thank you! Priya

[piechottam]

Hi,

thx for the details. Do you get the error if you run JACUSA without -b hs37d5.bed? Can you please share "hs37d5.bed"?

Please return with: java -jar JACUSA_v1.3.0.jar call-2 -a H:1 -p 26 -r rddsdec2chr1to22XY.out -s gDNAchr1to22andXY.bam TRNASTARwithMDchr1to22andXY.bam &> jacdec2chr1to22XYnooption.log

Is this the only Exception? [...] Exception in thread "Thread-21" java.lang.StackOverflowError [...]

Best, Michael

Am Di., 3. Dez. 2019 um 22:20 Uhr schrieb pswaminath < notifications@github.com>:

Hi Michael,

I ran JACUSA with the following command (complete bed file with positions in chr 1 to Y, bam file contains alignments chr 1 to Y)

Command used:

java -jar JACUSA_v1.3.0.jar call-2 -b hs37d5.bed -p 26 -r rddsdec2chr1to22XYNOaoption.out -s gDNAchr1to22andXY.bam TRNASTARwithMDchr1to22andXY.bam &> jacdec2chr1to22XYnooption.log

I have the complete log file, screenshot of tmp.gz files and the last lines of bed file attached here.

[image: tmpgzfilesscreenshot] https://user-images.githubusercontent.com/49890361/70089441-b19c7d00-15dd-11ea-9bc0-b7960f3f85bd.png jacdec2chr1to22XYNOaoption.log https://github.com/dieterich-lab/JACUSA/files/3918691/jacdec2chr1to22XYNOaoption.log

Please take a look.

I noticed that JACUSA run goes up to the positions in chr Y in the bed file provided by us and then it is stuck before wrapping up the result. Please confirm. It did not stop anywhere inbetween.

I have copied few lines where the error started and then the end of the messages. Please see attached log and screenshots for more information

[ INFO ] 05:19:44 : Started screening contig Y:28680525-28780524 [ INFO ] 05:19:44 : Started screening contig Y:58999981-59034049 Exception in thread "Thread-21" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

[image: hs37d5bedlastlinesscreenshot] https://user-images.githubusercontent.com/49890361/70090481-e3aede80-15df-11ea-98f9-ab9cacb5b0d2.png

Thank you! Priya

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[pswaminath]

Michael, I ran JACUSA initially with the complete bam file with no -b option including chr 1 to Y along with MT and other sequences - got error. I don't have the log file for this run. Those were java exceptions and the same message where it gets stuck at the last. at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446)

Yes I can share the bed file. Please see attached the txt file.

Exception in thread "Thread-21" java.lang.StackOverflowError . Yes this is the only exception I get without -a H:1 option

hs37d5.txt (I was not able to attach the bed file as is so I changed the file extension to .txt)

Thanks! Priya

[pswaminath]

Michael, I have the log file attached after your suggestion of running the command with -a H:1 and without bed file ( no -b option).

Command used:

java -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -p 26 -r rddsdec2chr1to22XYwithaoptionnob.out -s AVR085108gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec2chr1to22XYwithaoptionnob.log

I still get the error and it stops with the same message in the last after chromosome Y. There is an exception. See attached log file for complete details.

[ INFO ] 04:29:39 : Started screening contig Y:28680525-28780524 [ INFO ] 04:29:39 : Started screening contig Y:58999981-59099980 Exception in thread "Thread-20" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

jacdec2chr1to22XYwithaoptionnob.log

[piechottam]

Hi,

thx for the attached log. Okay, the problem is a library that we use to compute the test-statistic. Lib: org.apache.commons.math3 Method: digamma() The method is implemented as a recursion and is called very often -> Deep Recursion -> StackOverflowError!

I am not sure, why this is. Is your data on chromosome Y extremely covered? You could use "-D 1000" to limit the considered coverage per BAM and site to 1000 Reads.

Or, this is the preferred solution. You could increase the stack size of your JAVA virtual machine.

See here for a possible solution: https://stackoverflow.com/questions/3700459/how-to-increase-the-java-stack-size

java -Xss4M JACUSA....

Best, Michael

Am Mi., 4. Dez. 2019 um 23:13 Uhr schrieb pswaminath < notifications@github.com>:

Michael, I have the log file attached after your suggestion of running the command with -a H:1 and without bed file ( no -b option).

Command used:

java -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -p 26 -r rddsdec2chr1to22XYwithaoptionnob.out -s AVR085108gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec2chr1to22XYwithaoptionnob.log

I still get the error and it stops with the same message in the last after chromosome Y. There is an exception. See attached log file for complete details.

[ INFO ] 04:29:39 : Started screening contig Y:28680525-28780524 [ INFO ] 04:29:39 : Started screening contig Y:58999981-59099980 Exception in thread "Thread-20" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

jacdec2chr1to22XYwithaoptionnob.log https://github.com/dieterich-lab/JACUSA/files/3924139/jacdec2chr1to22XYwithaoptionnob.log

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[pswaminath]

Hi, Thanks for your suggestions. I will check running with -Xss option.

Regarding D option -D 1000 Can I check along with - a H:1 ? Does the command look like this then? java -Xss4m -jar JACUSA_v1.3.0.jar -a H:1 -D 1000 ........

Priya

[piechottam]

Hi, pls test only the -Xss Option. And filter your -b - pls retain only reads on chromosome Y, where the problems occur, right? This will save a lot of time.

Best, Michael

Am Do., 5. Dez. 2019 um 18:49 Uhr schrieb pswaminath < notifications@github.com>:

Hi, Thanks for your suggestions. I will check running with -Xss option.

Regarding D option -D 1000 Can I check along with - a H:1 ? Does the command look like this then? java -Xss4m -jar JACUSA_v1.3.0.jar -a H:1 -D 1000 ........

Priya

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[pswaminath]

Michael, Thanks for your suggestions. I checked chromosome Y only along with two other runs.

Here are my different runs and what I found. Please see the two error log files attached for the failed runs.

1) with Xss4m and -a H:1 but no b option - Same Error (after chromosome Y - Exception and stuck at the library) java -Xss4m -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -p 26 -r rddsdec5chr1to22XYwithaoptionnobXss.out -s AVR085108 gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec5chr1to22XYwithaoptionnobXss.log

Error lines

[ INFO ] 04:48:45 : Started screening contig Y:28680525-28780524 [ INFO ] 04:48:45 : Started screening contig Y:58999981-59099980 Exception in thread "Thread-23" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

2) with Xss4m, with b option for only chromsome Y in bed file it completed without any problem and no errors java -Xss4m -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -b hs37d5chrynew.bed -p 26 -r rddsdec5chr1to22XYwithaoptionwithbXssonlyY.out -s AVR085108gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec5chr1to22XYwithaoptionwithbXssonlyY.log

3)with Xss4m and -a H:1 and b with bed file (chr 1 to Y) java -Xss4m -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -b hs37d5.bed -p 26 -r rddsdec5chr1to22XYwithaoptionwithbfullXss.out -s AVR085108gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec5chr1to22XYwithaoptionwithbfullXss.log

Error lines [ INFO ] 05:10:06 : Started screening contig Y:28494610-28594609 [ INFO ] 05:10:06 : Started screening contig Y:28680525-28780524 [ INFO ] 05:10:06 : Started screening contig Y:58999981-59034049 Exception in thread "Thread-21" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

Log file for Run 1: jacdec5chr1to22XYwithaoptionnobXss.log

Log file for Run 3: jacdec5chr1to22XYwithaoptionwithbfullXss.log

Should I increase the stack size from 4m to a higher value? I am not sure what is the maximum should I keep with 26 threads? Will this help finish the run? Problem seems to happen at the last step where it is processing in the library in all the runs involving all the chromosomes (1 ... Y).

Thanks! Priya

[piechottam]

Hi Priya,

It is hard to come up with a suggestion for the correct value. maybe use Xss64m.

You could run each chromosome separately. At least you could pinpoint what chromosome is problematic.

Best, Michael

Am Fr., 6. Dez. 2019 um 17:18 Uhr schrieb pswaminath < notifications@github.com>:

Michael, Thanks for your suggestions. I checked chromosome Y only along with two other runs.

Here are my different runs and what I found. Please see the two error log files attached for the failed runs.

1. with Xss4m and -a H:1 but no b option - Same Error (after chromosome Y - Exception and stuck at the library) java -Xss4m -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -p 26 -r rddsdec5chr1to22XYwithaoptionnobXss.out -s AVR085108 gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec5chr1to22XYwithaoptionnobXss.log

Error lines

[ INFO ] 04:48:45 : Started screening contig Y:28680525-28780524 [ INFO ] 04:48:45 : Started screening contig Y:58999981-59099980 Exception in thread "Thread-23" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

1. with Xss4m, with b option for only chromsome Y in bed file it completed without any problem and no errors java -Xss4m -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -b hs37d5chrynew.bed -p 26 -r rddsdec5chr1to22XYwithaoptionwithbXssonlyY.out -s AVR085108gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec5chr1to22XYwithaoptionwithbXssonlyY.log

3)with Xss4m and -a H:1 and b with bed file (chr 1 to Y) java -Xss4m -Xmx50G -jar JACUSA_v1.3.0.jar call-2 -a H:1 -b hs37d5.bed -p 26 -r rddsdec5chr1to22XYwithaoptionwithbfullXss.out -s AVR085108gDNAchr1to22andXY.bam AVR085108TRNASTARwithMDchr1to22andXY.bam &> jacdec5chr1to22XYwithaoptionwithbfullXss.log

Error lines

[ INFO ] 05:10:06 : Started screening contig Y:28494610-28594609 [ INFO ] 05:10:06 : Started screening contig Y:28680525-28780524 [ INFO ] 05:10:06 : Started screening contig Y:58999981-59034049 Exception in thread "Thread-21" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:446) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461) at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

Log file for Run 1: jacdec5chr1to22XYwithaoptionnobXss.log https://github.com/dieterich-lab/JACUSA/files/3932937/jacdec5chr1to22XYwithaoptionnobXss.log

Log file for Run 3: jacdec5chr1to22XYwithaoptionwithbfullXss.log https://github.com/dieterich-lab/JACUSA/files/3933019/jacdec5chr1to22XYwithaoptionwithbfullXss.log

Should I increase the stack size from 4m to a higher value? I am not sure what is the maximum should I keep with 26 threads? Will this help finish the run? Problem seems to happen at the last step where it is processing in the library in all the runs involving all the chromosomes (1 ... Y).

Thanks! Priya

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[pswaminath]

Thanks Michael! I ran JACUSA with different stack sizes (Xss4m, Xmx10G and Xss20M,Xmx20G, Xss16M, Xmx25G. It did not work and stopped with StackOverflow error, same as before

Exception in thread "Thread-14" java.lang.StackOverflowError at org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)

After all the runs that I checked, the problem seem to occur mostly after finishing chromosome Y when wrapping up the result in the library is where it is stuck.

With Xss1024m, Xmx50G - I got Outofmemory Error.

I completed running few chromosomes seperately chr 1, 11, 12, 20 ,21, X, Y and chr20 and chr21 together. All the individual chromosome runs finished successfully. chr20 and chr21 also ran together successfully and got the output (stack size 8192KB).

When I ran chr 20, 21, 22, X and Y together it could not finish with the above stack size.

I have a run that is on with Xss64M and Xmx30G.

Priya

[piechottam]

Dear Priya,

I am confused.

Let me summarize your observations:

• Running each chromosome separately NO Exception
• Running Y with some other chromosome(s) gives you an Exception?

It is very hard to come up with a solution without looking at the data. You are the first to make such an observation. Does it help you if you run each analysis separately and combine the results afterwards?

One last option would be to reduce the number of parallel threads "-p

". You could try with 16 threads. Best, Michael Am Mi., 11. Dez. 2019 um 17:17 Uhr schrieb pswaminath < notifications@github.com>: > Thanks Michael! > I ran JACUSA with different stack sizes (Xss4m, Xmx10G and Xss20M,Xmx20G, > Xss16M, Xmx25G. It did not work and stopped with StackOverflow error, same > as before > > > *Exception in thread "Thread-14" java.lang.StackOverflowError at > org.apache.commons.math3.special.Gamma.digamma(Gamma.java:461)* > > After all the runs that I checked, the problem seem to occur mostly after > finishing chromosome Y when wrapping up the result in the library is where > it is stuck. > > *With Xss1024m, Xmx50G - I got Outofmemory Error.* > > I completed running few chromosomes seperately chr 1, 11, 12, 20 ,21, X, Y > and > chr20 and chr21 together. All the individual chromosome runs finished > successfully. chr20 and chr21 also ran together successfully and got the > output (stack size 8192KB). > > When I ran chr 20, 21, 22, X and Y together it could not finish with the > above stack size. > > I have a run that is on with Xss64M and Xmx30G. > > Priya > > — > You are receiving this because you commented. > Reply to this email directly, view it on GitHub > , > or unsubscribe > > . > -- Michael Piechotta Mob.:+49 176 623 828 74

[pswaminath]

Hi Michael, Thank you again! I am running the analysis and I will update you once done.

[pswaminath]

Hi Michael, Update regarding the analysis:

I ran JACUSA for each chromosome and then combined the results.

Thank you! Priya

[pswaminath]

Hi Michael, Regarding "Start" and "End" in the following line from analysis?

My questions:

I am in the process of annotating the variants. 1) Are these potential rna editing sites based on 1-based coordinate system? 2) Is the RNA editing taking place at position 13477 or at 13478? 3) how to understand base1 and base2 columns? G and AG (Is it a single nucleotide that is changing?)

"chrom" "chromStart" "chromEnd" "name" "stat" "strand" "base11" "bases21" "info" "filter_info" "cov1" "covs1" "cov2" "covs2" "cov" "matrix1.A" "matrix1.C" "matrix1.G" "matrix1.T" "base1" "matrix2.A" "matrix2.C" "matrix2.G" "matrix2.T" "base2" "baseChange" "editingFreq" "1" 13477 13478 "variant" 3.63902821722496 "." "0,0,36,0" "10,0,729,0" "" "" 36 36 739 739 775 0 0 36 0 "G" 10 0 729 0 "AG" "G->A" 0.013531799729364

Thank you! Priya

[piechottam]

Hi,

*1) Are these potential rna editing sites based on 1-based coordinate

system?*

0-based -> bed like file format

2) Is the RNA editing taking place at position 13477 or at 13478?

0-based: 13477 1-based: 13478

• 3) how to understand base1 and base2 columns? G and AG (Is it a single nucleotide that is changing?)*

"chrom" "chromStart" "chromEnd" "name" "stat" "strand" "base11" "bases21" "info" "filter_info" "cov1" "covs1" "cov2" "covs2" "cov" "matrix1.A" "matrix1.C" "matrix1.G" "matrix1.T" "base1" "matrix2.A" "matrix2.C" "matrix2.G" "matrix2.T" "base2" "baseChange" "editingFreq" "1" 13477 13478 "variant" 3.63902821722496 "." "0,0,36,0" "10,0,729,0" "" "" 36 36 739 739 775 0 0 36 0 "G" 10 0 729 0 "AG" "G->A" 0.013531799729364

Is this output from JACUSAhelper? This works for RNA-DNA-differences and assumes that that DNA was used as the first BAM file and RNA as the second in your JACUSA call.

AG are the observed bases. G->A the observed base change (DNA->RNA). In your DNA you have a G and in your RNA u might have A or AG.

Best, Michael

[pswaminath]

Michael, I appreciate you clarifying my questions. The output that I quoted is from JACUSAhelper.

You had mentioned: In your DNA you have a G and in your RNA u might have A or AG. Looking at base change column in the results, I thought that it was G->A only.

I am annotating these RNA editing sites using Annovar. Which one should I consider G->A or G->AG or both?

Thank you! Priya

[piechottam]

Hi, I hope I can clarify things. AG indicates observed bases per position G->A the observed base change. There is G in DNA and there is (maybe) G and A in RNA.

Best, Michael