error long spanning reads Malformed

Question: error long spanning reads Malformed

4.2 years ago by

Spain

Hi,

I have ran the Tophat 2.08 version and I have obtained this "result".log

[2014-09-10 09:47:28] Beginning TopHat run (v2.0.8b)
-----------------------------------------------
[2014-09-10 09:47:28] Checking for Bowtie
Bowtie version:   2.1.0.0
[2014-09-10 09:47:28] Checking for Samtools
Samtools version:   0.1.19.0
[2014-09-10 09:47:28] Checking for Bowtie index files
[2014-09-10 09:47:28] Checking for reference FASTA file
[2014-09-10 09:47:28] Generating SAM header for /home/xelcoro/Software/tophat-2.0.12.Linux_x86_64/mm10/mm10
format:   fastq
quality scale:   phred33 (default)
[2014-09-10 09:48:20] Reading known junctions from GTF file
[2014-09-10 09:48:24] Preparing reads
left reads: min. length=16, max. length=388, 133029181 kept reads (319 discarded)
Warning: short reads (<20bp) will make TopHat quite slow and take large amount of memory because they are likely to be mapped in too many places
[2014-09-10 10:32:23] Creating transcriptome data files..
[2014-09-10 10:33:01] Building Bowtie index from mm10.fa
[2014-09-10 10:45:05] Mapping left_kept_reads to transcriptome mm10 with Bowtie2
[2014-09-10 14:49:18] Resuming TopHat pipeline with unmapped reads
[2014-09-10 14:49:18] Mapping left_kept_reads.m2g_um to genome mm10 with Bowtie2
[2014-09-10 21:05:51] Mapping left_kept_reads.m2g_um_seg1 to genome mm10 with Bowtie2 (1/15)
[2014-09-10 23:20:33] Mapping left_kept_reads.m2g_um_seg2 to genome mm10 with Bowtie2 (2/15)
[2014-09-11 01:24:01] Mapping left_kept_reads.m2g_um_seg3 to genome mm10 with Bowtie2 (3/15)
[2014-09-11 03:36:15] Mapping left_kept_reads.m2g_um_seg4 to genome mm10 with Bowtie2 (4/15)
[2014-09-11 05:42:01] Mapping left_kept_reads.m2g_um_seg5 to genome mm10 with Bowtie2 (5/15)
[2014-09-11 07:31:05] Mapping left_kept_reads.m2g_um_seg6 to genome mm10 with Bowtie2 (6/15)
[2014-09-11 08:52:39] Mapping left_kept_reads.m2g_um_seg7 to genome mm10 with Bowtie2 (7/15)
[2014-09-11 09:30:02] Mapping left_kept_reads.m2g_um_seg8 to genome mm10 with Bowtie2 (8/15)
[2014-09-11 09:47:17] Mapping left_kept_reads.m2g_um_seg9 to genome mm10 with Bowtie2 (9/15)
[2014-09-11 09:53:55] Mapping left_kept_reads.m2g_um_seg10 to genome mm10 with Bowtie2 (10/15)
[2014-09-11 09:55:36] Mapping left_kept_reads.m2g_um_seg11 to genome mm10 with Bowtie2 (11/15)
[2014-09-11 09:55:48] Mapping left_kept_reads.m2g_um_seg12 to genome mm10 with Bowtie2 (12/15)
[2014-09-11 09:55:52] Mapping left_kept_reads.m2g_um_seg13 to genome mm10 with Bowtie2 (13/15)
[2014-09-11 09:55:56] Mapping left_kept_reads.m2g_um_seg14 to genome mm10 with Bowtie2 (14/15)
[2014-09-11 09:56:00] Mapping left_kept_reads.m2g_um_seg15 to genome mm10 with Bowtie2 (15/15)
[2014-09-11 09:56:04] Retrieving sequences for splices
[2014-09-11 10:00:03] Indexing splices
Building a SMALL index
[2014-09-11 10:00:28] Mapping left_kept_reads.m2g_um_seg1 to genome segment_juncs with Bowtie2 (1/15)
[2014-09-11 10:14:33] Mapping left_kept_reads.m2g_um_seg2 to genome segment_juncs with Bowtie2 (2/15)
[2014-09-11 10:27:41] Mapping left_kept_reads.m2g_um_seg3 to genome segment_juncs with Bowtie2 (3/15)
[2014-09-11 10:41:06] Mapping left_kept_reads.m2g_um_seg4 to genome segment_juncs with Bowtie2 (4/15)
[2014-09-11 10:53:45] Mapping left_kept_reads.m2g_um_seg5 to genome segment_juncs with Bowtie2 (5/15)
[2014-09-11 11:04:34] Mapping left_kept_reads.m2g_um_seg6 to genome segment_juncs with Bowtie2 (6/15)
[2014-09-11 11:12:10] Mapping left_kept_reads.m2g_um_seg7 to genome segment_juncs with Bowtie2 (7/15)
[2014-09-11 11:16:48] Mapping left_kept_reads.m2g_um_seg8 to genome segment_juncs with Bowtie2 (8/15)
[2014-09-11 11:19:08] Mapping left_kept_reads.m2g_um_seg9 to genome segment_juncs with Bowtie2 (9/15)
[2014-09-11 11:20:00] Mapping left_kept_reads.m2g_um_seg10 to genome segment_juncs with Bowtie2 (10/15)
[2014-09-11 11:20:17] Mapping left_kept_reads.m2g_um_seg11 to genome segment_juncs with Bowtie2 (11/15)
[2014-09-11 11:20:25] Mapping left_kept_reads.m2g_um_seg12 to genome segment_juncs with Bowtie2 (12/15)
[2014-09-11 11:20:33] Mapping left_kept_reads.m2g_um_seg13 to genome segment_juncs with Bowtie2 (13/15)
[2014-09-11 11:20:39] Mapping left_kept_reads.m2g_um_seg14 to genome segment_juncs with Bowtie2 (14/15)
[2014-09-11 11:20:45] Mapping left_kept_reads.m2g_um_seg15 to genome segment_juncs with Bowtie2 (15/15)
[2014-09-11 11:20:51] Joining segment hits
[FAILED]
Error running 'long_spanning_reads':Warning: 2476721 malformed closure

I am running 12 samples in a i3 computer with 16 Gb RAM.

What does this mean?

what should i do?

Thank you,

Xabier

rna-seq tophat alignment galaxy • 1.8k views

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modified 4.2 years ago by Jennifer Hillman Jackson ♦ 25k • written 4.2 years ago by xabierelcoro • 0

4.2 years ago by

Jennifer Hillman Jackson ♦ 25k

United States

Jennifer Hillman Jackson ♦ 25k wrote:

Hello,

I haven't come across this error myself, but from a bit of searching I found that many users have come across it. Most questions posted online do not have a remedy - but I did find one at seqanswers that suggests that the issue is associated with the reference annotation (GTF, GFF3) selection. The post is here:
http://seqanswers.com/forums/archive/index.php/t-30011.html

It sounds as if a mismatch between the reference annotation and the reference genome used in the run can trigger a problem, then this error can come up. The specific problem is unclear, but it could be an identifier mismatch issue. Meaning, the chromosome names are not an exact match between the two inputs causing a conflict when the junctions are reconciled to produce the output.

I would have guessed a different issue - not enough valid spliced hits, but I see no evidence of that being the specific cause. And it does not appear to be related to a memory issue (e.g. job being too large for the provided resources) from the given information.

Hopefully this helps! Jen, Galaxy team

ADD COMMENT • link written 4.2 years ago by Jennifer Hillman Jackson ♦ 25k

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