TopHat - Error: gtf_to_fasta returned an error.

[jthomas5062]

Hello all,

I am trying to perform alignments using TopHat2 and am receiving a gtf_to_fasta error message.

My command is the following:
$tophat -p 8 -G genes.gtf -o C1_R1_thout genome C1_R1_1.fq C1_R1_2.fq

The output:
[2014-07-28 15:40:04] Beginning TopHat run (v2.0.12)
-----------------------------------------------
[2014-07-28 15:40:04] Checking for Bowtie
Bowtie version: 2.2.3.0
[2014-07-28 15:40:05] Checking for Samtools
Samtools version: 0.1.19.0
[2014-07-28 15:40:05] Checking for Bowtie index files (genome)..
[2014-07-28 15:40:05] Checking for reference FASTA file
[2014-07-28 15:40:05] Generating SAM header for genome
[2014-07-28 15:40:05] Reading known junctions from GTF file
[2014-07-28 15:40:09] Preparing reads
left reads: min. length=75, max. length=75, 11607353 kept reads (0 discarded)
right reads: min. length=75, max. length=75, 11607353 kept reads (0 discarded)
[2014-07-28 15:45:43] Building transcriptome data files C1_R1_thout/tmp/genes
[FAILED]
Error: gtf_to_fasta returned an error.

When I look at the output error file "g2f.err", I see the following:
dyld: lazy symbol binding failed: Symbol not found: __ZNKSt5ctypeIcE13_M_widen_initEv
Referenced from: /usr/local/bin/gtf_to_fasta
Expected in: /usr/lib/libstdc++.6.dylib

dyld: Symbol not found: __ZNKSt5ctypeIcE13_M_widen_initEv
Referenced from: /usr/local/bin/gtf_to_fasta
Expected in: /usr/lib/libstdc++.6.dylib


Has anyone had similar problems? Thanks!
James

[jthomas5062]

The following files are in the directory from which I am executing the command:

genome.1.bt2
genome.2.bt2
genome.3.bt2
genome.4.bt2
genome.rev.1.bt2
genome.rev.2.bt2
genome.fa
genes.gtf
C1_R1_1.fq
C1_R1_2.fq

[GenoMax]

Did you download the Mac (this is a Mac I presume) binaries from TopHat site? Do you have Xcode installed on this Mac?

Are you running an older version of OS X? This *may* be a problem of trying to run the 64-bit binary on a 32-bit machine.

[jthomas5062]

I am running a 64-Bit Mac OS X 10.9.4; 16 GB RAM; 4 cores and Xcode is installed

[jthomas5062]

I'm using TopHat 2

Admins-iMac:~ jamesdthomas$ tophat -v
TopHat v2.0.12

[GenoMax]

Have you checked if the test data supplied with TopHat works without errors?

[jthomas5062]

My command:
tophat -p 8 -o test_thout test_ref reads_1.fq reads_2.fq

Output:
[2014-07-29 09:10:21] Beginning TopHat run (v2.0.12)
-----------------------------------------------
[2014-07-29 09:10:21] Checking for Bowtie
Bowtie version: 2.2.3.0
[2014-07-29 09:10:21] Checking for Samtools
Samtools version: 0.1.19.0
[2014-07-29 09:10:21] Checking for Bowtie index files (genome)..
[2014-07-29 09:10:21] Checking for reference FASTA file
[2014-07-29 09:10:21] Generating SAM header for test_ref
[2014-07-29 09:10:21] Preparing reads
left reads: min. length=75, max. length=75, 100 kept reads (0 discarded)
right reads: min. length=75, max. length=75, 100 kept reads (0 discarded)
[2014-07-29 09:10:21] Mapping left_kept_reads to genome test_ref with Bowtie2
[2014-07-29 09:10:21] Mapping left_kept_reads_seg1 to genome test_ref with Bowtie2 (1/3)
[2014-07-29 09:10:22] Mapping left_kept_reads_seg2 to genome test_ref with Bowtie2 (2/3)
[2014-07-29 09:10:22] Mapping left_kept_reads_seg3 to genome test_ref with Bowtie2 (3/3)
[2014-07-29 09:10:22] Mapping right_kept_reads to genome test_ref with Bowtie2
[2014-07-29 09:10:22] Mapping right_kept_reads_seg1 to genome test_ref with Bowtie2 (1/3)
[2014-07-29 09:10:22] Mapping right_kept_reads_seg2 to genome test_ref with Bowtie2 (2/3)
[2014-07-29 09:10:22] Mapping right_kept_reads_seg3 to genome test_ref with Bowtie2 (3/3)
[2014-07-29 09:10:22] Searching for junctions via segment mapping
[FAILED]
Error: segment-based junction search failed with err =-5


When I look in segment_juncs log, I see the same error as with my actual dataset:
segment_juncs v2.0.12 (4277)
---------------------------
[samopen] SAM header is present: 1 sequences.
Loading reference sequences...
Loading test_chromosome...done
Loading ...done
>> Performing segment-search:
Loading left segment hits...
dyld: lazy symbol binding failed: Symbol not found: __ZNKSt5ctypeIcE13_M_widen_initEv
Referenced from: /usr/local/bin/segment_juncs
Expected in: /usr/lib/libstdc++.6.dylib

dyld: Symbol not found: __ZNKSt5ctypeIcE13_M_widen_initEv
Referenced from: /usr/local/bin/segment_juncs
Expected in: /usr/lib/libstdc++.6.dylib


It seems like more of a "my computer problem" than a TopHat specific problem. I'm somewhat of a novice when it comes to libraries, etc. When I tried to search for a solution to these "symbol not found" errors, I came up pretty empty handed.

Thanks,
James

[GenoMax]

Can you try a previous version of TopHat with the test data (say v.2.0.11, http://ccb.jhu.edu/software/tophat/downloads/) to see if this is a problem with the latest Mac binary that is being supplied?

[jthomas5062]

I downloaded v.2.0.11 of TopHat and am now getting this error message after running on the test data set:

[2014-07-30 16:36:24] Beginning TopHat run (v2.0.11)
-----------------------------------------------
[2014-07-30 16:36:24] Checking for Bowtie
Bowtie version: 2.2.3.0
[2014-07-30 16:36:24] Checking for Samtools
Samtools version: 0.1.19.0
[2014-07-30 16:36:24] Checking for Bowtie index files (genome)..
[2014-07-30 16:36:24] Checking for reference FASTA file
[2014-07-30 16:36:24] Generating SAM header for test_ref
[2014-07-30 16:36:24] Preparing reads
left reads: min. length=75, max. length=75, 100 kept reads (0 discarded)
right reads: min. length=75, max. length=75, 100 kept reads (0 discarded)
[2014-07-30 16:36:24] Mapping left_kept_reads to genome test_ref with Bowtie2
[FAILED]
Error running:
/usr/local/bin/bam2fastx --all testdata_thout/tmp/left_kept_reads.bam|/usr/local/bin/bowtie2 -k 20 -D 15 -R 2 -N 0 -L 20 -i S,1,1.25 --gbar 4 --mp 6,2 --np 1 --rdg 5,3 --rfg 5,3 --score-min C,-14,0 -p 8 --sam-no-hd -x test_ref -|/usr/local/bin/fix_map_ordering --bowtie2-min-score 15 --read-mismatches 2 --read-gap-length 2 --read-edit-dist 2 --read-realign-edit-dist 3 --index-outfile testdata_thout/tmp/left_kept_reads.mapped.bam.index --sam-header testdata_thout/tmp/test_ref_genome.bwt.samheader.sam - testdata_thout/tmp/left_kept_reads.mapped.bam testdata_thout/tmp/left_kept_reads_unmapped.bam

[jthomas5062]

My submmitted commands:

mkdir testdata_thout

tophat2 -p 8 -o testdata_thout test_ref reads_1.fq reads_2.fq

[zhangxw]

TopHat - Error: gtf_to_fasta returned an error.

has this problem solved? I met the same issue.

[GenoMax]

has this problem solved? I met the same issue.

Just to be specific. You are using a Mac and have had the exact same issue as jthomas5062.

With the test data and/or your own?

[zhangxw]

Yes, I am using Mac OS.
Same problem with the test_data by TopHat run (v2.0.11)

[zhangxw]

Such problem disappears with the test_data by TapHat v2.0.10;

however, in TapHat v2.0.10; it has Error: gtf_to_fasta returned an error. with my own data.

[GenoMax]

It appears that all Mac binaries since 2.0.10 have this problem. -stdlib option to build the binary either needs libstdc++ or libc++ (am not a developer) and it does not appear to be properly linked for Mac binaries. I am surprised that no one has reported this problem to the developers. Until the authors correct this we may be out-of-luck.

Has anyone tried to compile from source?

The last version of TopHat that is functional on OS X (10.9.x) is v.2.0.9.