TopHat err = -9

[DineshCyanam]

I am trying to run my RNA-seq data in TopHat and I keep getting the below error. I have tried the latest 1.1.0 version and it gave me the same error. So I went back to 1.0.14 and I got output for 1 sample. Now using 1.0.14, I tried to run another sample but I am getting the same error.

tophat --num-threads 8 -r 80 --output-dir /home/dinesh/Samples/171/tophat_out_67625/ /home/dinesh/hg19/hg19 s_4_1_sequence.fq,s_7_1_sequence.fq s_4_2_sequence.fq,s_7_2_sequence.fq

[Mon Oct 11 00:44:32 2010] Beginning TopHat run (v1.0.14)
-----------------------------------------------
[Mon Oct 11 00:44:32 2010] Preparing output location /home/dinesh/Samples/171/tophat_out_67625//
[Mon Oct 11 00:44:32 2010] Checking for Bowtie index files
[Mon Oct 11 00:44:32 2010] Checking for reference FASTA file
[Mon Oct 11 00:44:32 2010] Checking for Bowtie
	Bowtie version:		 0.12.7.0
[Mon Oct 11 00:44:32 2010] Checking reads
	seed length:	 60bp
	format:		 fastq
	quality scale:	 phred33 (default)
[Mon Oct 11 01:20:03 2010] Mapping reads against hg19 with Bowtie
[Mon Oct 11 01:56:26 2010] Joining segment hits
	Splitting reads into 2 segments
[Mon Oct 11 02:11:51 2010] Mapping reads against hg19 with Bowtie
[Mon Oct 11 03:25:32 2010] Mapping reads against hg19 with Bowtie
[Mon Oct 11 04:40:38 2010] Mapping reads against hg19 with Bowtie
[Mon Oct 11 05:14:31 2010] Joining segment hits
	Splitting reads into 2 segments
[Mon Oct 11 05:30:01 2010] Mapping reads against hg19 with Bowtie
[Mon Oct 11 06:39:51 2010] Mapping reads against hg19 with Bowtie
[Mon Oct 11 07:45:55 2010] Searching for junctions via segment mapping
	[FAILED]
Error: segment-based junction search failed with err = -9

I have also mailed Cole Trapnell the error and the log files and haven't heard from him yet. Any insight into this error is greatly appreciated.

[plabaj]

I can only say that I have the same problem and no idea what is wrong


[Sat Oct 9 18:42:58 2010] Beginning TopHat run (v1.1.0)
-----------------------------------------------
[Sat Oct 9 18:42:58 2010] Preparing output location 2_rep/
[Sat Oct 9 18:42:58 2010] Checking for Bowtie index files
[Sat Oct 9 18:42:58 2010] Checking for reference FASTA file
[Sat Oct 9 18:42:58 2010] Checking for Bowtie
Bowtie version: 0.12.7.0
[Sat Oct 9 18:42:58 2010] Checking for Samtools
Samtools version: 0.1.8.0
[Sat Oct 9 18:43:00 2010] Checking reads
min read length: 50bp, max read length: 50bp
format: fasta
[Sat Oct 9 21:39:25 2010] Mapping reads against hg19_c with Bowtie
[Sun Oct 10 06:34:25 2010] Joining segment hits
[Sun Oct 10 10:34:55 2010] Mapping reads against hg19_c with Bowtie(1/2)
[Sun Oct 10 16:55:39 2010] Mapping reads against hg19_c with Bowtie(2/2)
[Sun Oct 10 22:24:52 2010] Searching for junctions via segment mapping
[FAILED]
Error: segment-based junction search failed with err = -9

Could it be some memory issue?
I was running 3 instances on the one machine and now I can see that two others, which are still running, are using 28-30GB of RAM.
Machine has only 72, so if the third instance came when two first have taken everything what they need it could happened that there was no more for the third one.

I've currently restarted this instance on a separate machine.
I'll let you know how it went.

[DineshCyanam]

Well I am running just 1 instance of TopHat on our super computing cluster with 8 threads. So I don't think memory is a problem. Anyways I will talk to the sys ops and find out how much memory TopHat was using.

But what I don't understand is that TopHat was designed to run on a normal desktop environment. Here is an excerpt from the paper "TopHat: discovering splice junctions with RNA-Seq" (PMID:19289445)

The TopHat pipeline is much faster than previous systems, mapping nearly 2.2 million reads per CPU hour, which is sufficient to process an entire RNA-Seq experiment in less than a day on a standard desktop computer.

Looking forward for your findings, Plabaj

- Dinesh

[plabaj]

After restarting I haven't noticed such error any more.
So I have no idea what was that.

Best,
Pawel Labaj

[DineshCyanam]

Plabaj, Did you figure out what the problem was? I am still encountering the same error.

[plabaj]

Unfortunately not.
Everything is working fine so far.

[zorph]

I seem to be having the same problem. Did you ever resolve this problem and if so, how?

Thanks in advance.

[bcalder]

This is a memory issue. Since we moved to doing Hiseq directional RNA-seq we've been having to reserve special nodes for processing these jobs (>32GB). And forget paired end... It gets to the segment_juncs step, goes single threaded and chews up all the memory.

Hopefully the authors have become aware of the problem.

[milesgr]

Has anyone resolved this issue with regards to paired-end colorspace data?

[diya]

Error message while running tophat

Hi,

I am trying to run tophat for the RNA-Seq paired end data. I have the following error message


[Fri Feb 10 11:35:13 2012] Beginning TopHat run (v1.2.0)
-----------------------------------------------
[Fri Feb 10 11:35:13 2012] Preparing output location ./tophat_out/
[Fri Feb 10 11:35:13 2012] Checking for Bowtie index files
[Fri Feb 10 11:35:13 2012] Checking for reference FASTA file
[Fri Feb 10 11:35:13 2012] Checking for Bowtie
Bowtie version: 0.12.7.0
[Fri Feb 10 11:35:13 2012] Checking for Samtools
Samtools Version: 0.1.10
[Fri Feb 10 11:35:51 2012] Checking reads
min read length: 80bp, max read length: 80bp
format: fastq
quality scale: phred33 (default)
[Fri Feb 10 12:00:19 2012] Mapping reads against hg19 with Bowtie
[Fri Feb 10 13:43:36 2012] Joining segment hits
[Fri Feb 10 13:50:56 2012] Mapping reads against hg19 with Bowtie(1/3)
[Fri Feb 10 16:04:25 2012] Mapping reads against hg19 with Bowtie(2/3)
[Fri Feb 10 18:20:29 2012] Mapping reads against hg19 with Bowtie(3/3)
[Fri Feb 10 19:30:34 2012] Mapping reads against hg19 with Bowtie
[Fri Feb 10 21:18:30 2012] Joining segment hits
[Fri Feb 10 21:26:32 2012] Mapping reads against hg19 with Bowtie(1/3)
[Sat Feb 11 00:24:15 2012] Mapping reads against hg19 with Bowtie(2/3)
[Sat Feb 11 03:26:59 2012] Mapping reads against hg19 with Bowtie(3/3)
[Sat Feb 11 05:02:57 2012] Searching for junctions via segment mapping
[FAILED]
Error: segment-based junction search failed with err = -11


Any help on this is greatly appreciated

[DineshCyanam]

Hi Diya,
Can you please post the command that you are using to run Tophat. Also, the Tophat version you are using is 1.2.0 which is more than an year old and there have been lots of changes since then. Please use the latest version found at http://tophat.cbcb.umd.edu