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tophat mapping bug? cedance Bioinformatics 2 11-11-2011 08:56 AM
TopHat 1.3.0 Splicing Bug ? Dario1984 Bioinformatics 11 09-16-2011 09:15 PM
bug for the tophat 1.3 beta liuxq Bioinformatics 4 06-09-2011 11:25 AM
TopHat bug telos Bioinformatics 0 03-17-2010 08:50 AM
Tophat Bug? jling Bioinformatics 0 02-11-2010 04:14 PM

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Old 06-02-2011, 06:17 PM   #1
peromhc
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Default tophat 1.3.0 bug??

any ideas?


Code:
matthew@macmanes:~/tophat-1.3.0$ /home/matthew/tophat-1.3.0/tophat -o /media/hd/annotation/tophat/solitary -r 100 -p 16 /media/hd/annotation/tophat/bowtie.index/tuco.mrna.annotate.2June11 /media/hd/tuco/mrna/peruanus/clipped/cp.final.clip_1.fq /media/hd/tuco/mrna/peruanus/clipped/cp.final.clip_2.fq

[Thu Jun  2 17:47:45 2011] Beginning TopHat run (v1.3.0)
-----------------------------------------------
[Thu Jun  2 17:47:45 2011] Preparing output location /media/hd/annotation/tophat/solitary/
[Thu Jun  2 17:47:45 2011] Checking for Bowtie index files
[Thu Jun  2 17:47:45 2011] Checking for reference FASTA file
[Thu Jun  2 17:47:45 2011] Checking for Bowtie
	Bowtie version:			 0.12.7.0
[Thu Jun  2 17:47:45 2011] Checking for Samtools
	Samtools Version: 0.1.16
[Thu Jun  2 17:47:45 2011] Generating SAM header for /media/hd/annotation/tophat/bowtie.index/tuco.mrna.annotate.2June11
[Thu Jun  2 17:47:49 2011] Preparing reads
	format:		 fastq
	quality scale:	 phred33 (default)
Traceback (most recent call last):
 File "/home/matthew/tophat-1.3.0/tophat", line 2584, in <module>
   sys.exit(main())
 File "/home/matthew/tophat-1.3.0/tophat", line 2511, in main
   "left_kept_reads", "Left ")
 File "/home/matthew/tophat-1.3.0/tophat", line 1362, in prep_reads
   shell_cmd = ' '.join(filter_cmd)
TypeError: sequence item 50: expected string, NoneType found
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Old 06-02-2011, 07:06 PM   #2
liuxq
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get the same error
Quote:
Originally Posted by peromhc View Post
any ideas?


Code:
matthew@macmanes:~/tophat-1.3.0$ /home/matthew/tophat-1.3.0/tophat -o /media/hd/annotation/tophat/solitary -r 100 -p 16 /media/hd/annotation/tophat/bowtie.index/tuco.mrna.annotate.2June11 /media/hd/tuco/mrna/peruanus/clipped/cp.final.clip_1.fq /media/hd/tuco/mrna/peruanus/clipped/cp.final.clip_2.fq

[Thu Jun  2 17:47:45 2011] Beginning TopHat run (v1.3.0)
-----------------------------------------------
[Thu Jun  2 17:47:45 2011] Preparing output location /media/hd/annotation/tophat/solitary/
[Thu Jun  2 17:47:45 2011] Checking for Bowtie index files
[Thu Jun  2 17:47:45 2011] Checking for reference FASTA file
[Thu Jun  2 17:47:45 2011] Checking for Bowtie
	Bowtie version:			 0.12.7.0
[Thu Jun  2 17:47:45 2011] Checking for Samtools
	Samtools Version: 0.1.16
[Thu Jun  2 17:47:45 2011] Generating SAM header for /media/hd/annotation/tophat/bowtie.index/tuco.mrna.annotate.2June11
[Thu Jun  2 17:47:49 2011] Preparing reads
	format:		 fastq
	quality scale:	 phred33 (default)
Traceback (most recent call last):
 File "/home/matthew/tophat-1.3.0/tophat", line 2584, in <module>
   sys.exit(main())
 File "/home/matthew/tophat-1.3.0/tophat", line 2511, in main
   "left_kept_reads", "Left ")
 File "/home/matthew/tophat-1.3.0/tophat", line 1362, in prep_reads
   shell_cmd = ' '.join(filter_cmd)
TypeError: sequence item 50: expected string, NoneType found
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Old 06-02-2011, 08:07 PM   #3
jmartin127
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Yep, same for me.
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Old 06-02-2011, 10:00 PM   #4
Dario1984
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I have another issue about the bowtie indexes :

Code:
[Fri Jun  3 14:29:02 2011] Beginning TopHat run (v1.3.0)
-----------------------------------------------
[Fri Jun  3 14:29:02 2011] Preparing output location /home/darstr/tmp/rnaSeqTH1.3/P/
[Fri Jun  3 14:29:02 2011] Checking for Bowtie index files
Error: Could not find Bowtie index files hg18.*
darstr@clark-lab:~/tmp$ echo $BOWTIE_INDEXES
/usr/local/share/indexes/hg18
darstr@clark-lab:~/tmp$ ls $BOWTIE_INDEXES
hg18.1.ebwt  hg18.2.ebwt  hg18.3.ebwt  hg18.4.ebwt  hg18.rev.1.ebwt  hg18.rev.2.ebwt
But the user manual clearly says "... then looks in the directory specified in the BOWTIE_INDEXES environment variable."

Also, does anyone send questions or bug reports to the official tophat.cufflinks@gmail.com address ? It doesn't seem to be monitored.
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Old 06-03-2011, 01:28 AM   #5
liuxq
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new bug
tophat --no-novel-juncs --phred64-quals -G ensembl_refGene_ucscGene.gtf -o output genome_index/bowtie/mm9/mm9 s_3_sequence.txt
Quote:
Traceback (most recent call last):
File "/home/liuxq/mybin/tools/tophat", line 2584, in <module>
sys.exit(main())
File "/home/liuxq/mybin/tools/tophat", line 2543, in main
user_supplied_deletions)
File "/home/liuxq/mybin/tools/tophat", line 2376, in spliced_alignment
os.remove(unspl_bwtfile)
UnboundLocalError: local variable 'unspl_bwtfile' referenced before assignment
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Old 06-03-2011, 01:35 AM   #6
liuxq
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what's your tophat command? use "hg18" for the <bowtie_index> parameter
Quote:
Originally Posted by Dario1984 View Post
I have another issue about the bowtie indexes :

Code:
[Fri Jun  3 14:29:02 2011] Beginning TopHat run (v1.3.0)
-----------------------------------------------
[Fri Jun  3 14:29:02 2011] Preparing output location /home/darstr/tmp/rnaSeqTH1.3/P/
[Fri Jun  3 14:29:02 2011] Checking for Bowtie index files
Error: Could not find Bowtie index files hg18.*
darstr@clark-lab:~/tmp$ echo $BOWTIE_INDEXES
/usr/local/share/indexes/hg18
darstr@clark-lab:~/tmp$ ls $BOWTIE_INDEXES
hg18.1.ebwt  hg18.2.ebwt  hg18.3.ebwt  hg18.4.ebwt  hg18.rev.1.ebwt  hg18.rev.2.ebwt
But the user manual clearly says "... then looks in the directory specified in the BOWTIE_INDEXES environment variable."

Also, does anyone send questions or bug reports to the official tophat.cufflinks@gmail.com address ? It doesn't seem to be monitored.
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Old 06-03-2011, 06:17 AM   #7
Cole Trapnell
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Quote:
Originally Posted by peromhc View Post
any ideas?


Code:
matthew@macmanes:~/tophat-1.3.0$ /home/matthew/tophat-1.3.0/tophat -o /media/hd/annotation/tophat/solitary -r 100 -p 16 /media/hd/annotation/tophat/bowtie.index/tuco.mrna.annotate.2June11 /media/hd/tuco/mrna/peruanus/clipped/cp.final.clip_1.fq /media/hd/tuco/mrna/peruanus/clipped/cp.final.clip_2.fq

[Thu Jun  2 17:47:45 2011] Beginning TopHat run (v1.3.0)
-----------------------------------------------
[Thu Jun  2 17:47:45 2011] Preparing output location /media/hd/annotation/tophat/solitary/
[Thu Jun  2 17:47:45 2011] Checking for Bowtie index files
[Thu Jun  2 17:47:45 2011] Checking for reference FASTA file
[Thu Jun  2 17:47:45 2011] Checking for Bowtie
	Bowtie version:			 0.12.7.0
[Thu Jun  2 17:47:45 2011] Checking for Samtools
	Samtools Version: 0.1.16
[Thu Jun  2 17:47:45 2011] Generating SAM header for /media/hd/annotation/tophat/bowtie.index/tuco.mrna.annotate.2June11
[Thu Jun  2 17:47:49 2011] Preparing reads
	format:		 fastq
	quality scale:	 phred33 (default)
Traceback (most recent call last):
 File "/home/matthew/tophat-1.3.0/tophat", line 2584, in <module>
   sys.exit(main())
 File "/home/matthew/tophat-1.3.0/tophat", line 2511, in main
   "left_kept_reads", "Left ")
 File "/home/matthew/tophat-1.3.0/tophat", line 1362, in prep_reads
   shell_cmd = ' '.join(filter_cmd)
TypeError: sequence item 50: expected string, NoneType found

This should now be fixed. We pushed an update to the 1.3.0 tarballs late last night.
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Old 06-03-2011, 08:31 AM   #8
jmartin127
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Thanks Cole for the quick fix. Version 1.3.0 is running successfully now.

Quote:
Originally Posted by Cole Trapnell View Post
This should now be fixed. We pushed an update to the 1.3.0 tarballs late last night.
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Old 06-03-2011, 08:00 PM   #9
Dario1984
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Quote:
Originally Posted by liuxq View Post
what's your tophat command? use "hg18" for the <bowtie_index> parameter
Yeah, I am using that. Also, it works if I cd into the directory where the indexes are. It just doesn't work from outside the directory, when the environment variable set to the path of the indexes, as the TopHat documentation says to do.
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Old 06-06-2011, 07:29 AM   #10
berath
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I also get the same error message in post#5. an update on the binaries would be much appreciated.
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Old 06-07-2011, 02:32 PM   #11
Auction
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Quote:
Originally Posted by berath View Post
I also get the same error message in post#5. an update on the binaries would be much appreciated.
I also got this error message. My parameter is
-p 8 -g 1 -G Mus_musculus.NCBIM37.62.gtf.new --no-novel-juncs --no-novel-indels --solexa1.3-qual
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Old 06-08-2011, 08:40 AM   #12
babbitt
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I have a different error message. This was using tophat 1.3.0 on paired-end solid reads, which had been successfully mapped using tophat 1.2.0. The error message reads:

Error: qual length (57) differs from seq length (51) for fastq record 83_1325_28_F3!
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Old 06-09-2011, 03:15 AM   #13
ramu
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Default bug fix

Quote:
Originally Posted by liuxq View Post
new bug
tophat --no-novel-juncs --phred64-quals -G ensembl_refGene_ucscGene.gtf -o output genome_index/bowtie/mm9/mm9 s_3_sequence.txt

Here is a fix for
"UnboundLocalError: local variable 'unspl_samfile' referenced before assignment"
error.

Edit tophat and comment out ( usually it is in /usr/bin/tophat )
#os.remove(unspl_bwtfile) #os.remove(unspl_samfile)
my_dummy = 0 # to keep the if statment intact

The real fix should come from Cole!

Ramu
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Old 06-09-2011, 09:24 AM   #14
cwng
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Quote:
Originally Posted by ramu View Post
Here is a fix for
"UnboundLocalError: local variable 'unspl_samfile' referenced before assignment"
error.

Edit tophat and comment out ( usually it is in /usr/bin/tophat )
#os.remove(unspl_bwtfile) #os.remove(unspl_samfile)
my_dummy = 0 # to keep the if statment intact

The real fix should come from Cole!

Ramu
Another work around is to specify the "--keep-tmp" option to keep temporary files. The error is associated with the source code attempting to delete a temporary file before it is referenced. After the run is finished, you can delete the "tmp/" subdirectory.

Or just wait for the actual fix

-C
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Old 06-10-2011, 08:18 AM   #15
jbrwn
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Quote:
Originally Posted by babbitt View Post
I have a different error message. This was using tophat 1.3.0 on paired-end solid reads, which had been successfully mapped using tophat 1.2.0. The error message reads:

Error: qual length (57) differs from seq length (51) for fastq record 83_1325_28_F3!
I also receive this error when running pair-end reads from the SOLiD. This error shows up in the segment_juncs.log

Code:
segment_juncs v1.3.0 (2398:2399)
---------------------------
Loading reference sequences...
	Loading chr1...done
	Loading chr2...done
	Loading chr3...done
	Loading chr4...done
	Loading chr5...done
	Loading chr6...done
	Loading chr7...done
	Loading chr8...done
	Loading chr9...done
	Loading chr10...done
	Loading chr11...done
	Loading chr12...done
	Loading chr13...done
	Loading chr14...done
	Loading chr15...done
	Loading chr16...done
	Loading chr17...done
	Loading chr18...done
	Loading chr19...done
	Loading chr20...done
	Loading chr21...done
	Loading chr22...done
	Loading chrX...done
	Loading chrY...done
Loading left segment hits...
Error: qual length (56) differs from seq length (51) for fastq record 9_274_760_F3!
Edit: I should mention that doing a grep on the csfasta and qual shows seq length to be 51 (due to colorspace) with a qual length of 50.

That is all after receiving "err =1" from Tophat's stdout:
Code:
[Thu Jun  9 13:48:18 2011] Preparing reads
	format:		 fasta
	Left  reads: min. length=51, count=151286332
	Right reads: min. length=36, count=151112639
[Thu Jun  9 15:33:59 2011] Mapping left_kept_reads against hg19All.fa with Bowtie 
[Thu Jun  9 16:49:23 2011] Processing bowtie hits
[Thu Jun  9 18:31:50 2011] Mapping left_kept_reads_seg1 against hg19All.fa with Bowtie (1/2)
[Thu Jun  9 21:19:13 2011] Mapping left_kept_reads_seg2 against hg19All.fa with Bowtie (2/2)
[Fri Jun 10 00:11:09 2011] Mapping right_kept_reads against hg19All.fa with Bowtie 
[Fri Jun 10 01:26:25 2011] Processing bowtie hits
[Fri Jun 10 02:40:22 2011] Mapping right_kept_reads_seg1 against hg19All.fa with Bowtie (1/1)
[Fri Jun 10 03:39:36 2011] Searching for junctions via segment mapping
	[FAILED]
Error: segment-based junction search failed with err =1
Using these options:
Code:
tophat --color --quals --mate-inner-dist 200 --num-threads 8 --tmp-dir $tmp --output-dir $out --library-type fr-secondstrand $colorspaceindex $f3read $f5read $f3qual $f5qual

Last edited by jbrwn; 06-10-2011 at 08:25 AM.
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Old 06-16-2011, 08:36 AM   #16
ramu
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Default cuffmerge badly broken

Cufflinks v1.0.3, I get
__________________________________________________________
You are using Cufflinks v1.0.3, which is the most recent release.
[bam_header_read] EOF marker is absent. The input is probably truncated.
[bam_header_read] invalid BAM binary header (this is not a BAM file).
File allMerged_refseq/tmp/mergeSam_filenInW01 doesn't appear to be a valid BAM file, trying SAM...
[16:10:58] Loading reference annotation.
[16:11:00] Inspecting reads and determining fragment length distribution.

Error: this SAM file doesn't appear to be correctly sorted!
current hit is at Zv9_NA110:3369, last one was at chrM:11038
Cufflinks requires that if your file has SQ records in
the SAM header that they appear in the same order as the chromosomes names
in the alignments.
If there are no SQ records in the header, or if the header is missing,
the alignments must be sorted lexicographically by chromsome
name and by position.

[FAILED]
_______________________________________________________________

I had to edit the merged_blabla and re-arrange the chromosome order and run
the commands from the log file.

Any fix?
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Old 06-18-2011, 04:12 AM   #17
Fender_b
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Hi, I have the error: Error: segment-based junction search failed with err =1 when searching for junctions via segment mapping.

My log file says: Error: qual length (55) differs from seq length (51) for fastq record 427_275_933_F3!

Anyone has any fix/idea?

Thank you!
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