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DaehwanKimLab / tophat

Spliced read mapper for RNA-Seq
http://ccb.jhu.edu/software/tophat
Boost Software License 1.0
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Tophat 2.1.1 not compatible with bowtie2 v2.3 #43

Open rosbashlab opened 7 years ago

rosbashlab commented 7 years ago

Tophat 2.1.1 was not compatible with bowtie2 v2.3 in our server. To get tophat to work, I had to switch to bowtie2 v2.2.9.

I was getting this error message with bowtie2 v2.3

[2017-02-13 19:42:59] Checking for Bowtie Bowtie version: 2.3.0.0 [2017-02-13 19:42:59] Checking for Bowtie index files (genome).. Found both Bowtie1 and Bowtie2 indexes. [2017-02-13 19:42:59] Checking for reference FASTA file [2017-02-13 19:42:59] Generating SAM header for /home/analysis/genome/dm3/Sequence/bowtie_indexes/genome [2017-02-13 19:42:59] Reading known junctions from GTF file [2017-02-13 19:43:00] Preparing reads left reads: min. length=51, max. length=51, 8438399 kept reads (500 discarded) [2017-02-13 19:44:15] Building transcriptome data files WT_ZT06.tophat.out/tmp/genes [2017-02-13 19:44:17] Building Bowtie index from genes.fa [2017-02-13 19:51:06] Mapping left_kept_reads to transcriptome genes with Bowtie2 [2017-02-13 19:53:18] Resuming TopHat pipeline with unmapped reads [2017-02-13 19:53:18] Mapping left_kept_reads.m2g_um to genome genome with Bowtie2 [FAILED] Error running bowtie: Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error while flushing and closing output Error: Encountered exception: 'Unidentified exception' Command: /opt/BOWTIE2/2.3/bin/bowtie2-align-s --wrapper basic-0 -k 2 -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 10 --sam-no-hd -x /home/analysis/genome/dm3/Sequence/bowtie_indexes/genome - (ERR): bowtie2-align exited with value 1

nieder commented 7 years ago

The tophat2-2.1.1 test suite fails if bowtie-2.3.0 is installed:

cd ../test_data
/sw/bin/python2.7 /sw/build.build/tophat2-2.1.1-3/tophat-2.1.1/src/tophat -r 20 test_ref reads_1.fq reads_2.fq || exit 2

[2017-04-02 07:17:40] Beginning TopHat run (v2.1.1)
-----------------------------------------------
[2017-04-02 07:17:40] Checking for Bowtie
          Bowtie version:    2.3.0.0
[2017-04-02 07:17:40] Checking for Bowtie index files (genome)..
    Found both Bowtie1 and Bowtie2 indexes.
[2017-04-02 07:17:40] Checking for reference FASTA file
[2017-04-02 07:17:40] Generating SAM header for test_ref
[2017-04-02 07:17:40] 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)
[2017-04-02 07:17:40] Mapping left_kept_reads to genome test_ref with Bowtie2 
    [FAILED]
Error running:
/sw/build.build/tophat2-2.1.1-3/tophat-2.1.1/src/bam2fastx --all ./tophat_out/tmp/left_kept_reads.bam|/sw/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 1 --sam-no-hd -x test_ref -|/sw/build.build/tophat2-2.1.1-3/tophat-2.1.1/src/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 ./tophat_out/tmp/left_kept_reads.mapped.bam.index --sam-header ./tophat_out/tmp/test_ref_genome.bwt.samheader.sam - ./tophat_out/tmp/left_kept_reads.mapped.bam ./tophat_out/tmp/left_kept_reads_unmapped.bam

If using bowtie-2.2.9, then the test suite passes:

cd ../test_data
/sw/bin/python2.7 /sw/build.build/tophat2-2.1.1-3/tophat-2.1.1/src/tophat -r 20 test_ref reads_1.fq reads_2.fq || exit 2

[2017-04-03 07:22:56] Beginning TopHat run (v2.1.1)
-----------------------------------------------
[2017-04-03 07:22:56] Checking for Bowtie
          Bowtie version:    2.2.9.0
[2017-04-03 07:22:56] Checking for Bowtie index files (genome)..
    Found both Bowtie1 and Bowtie2 indexes.
[2017-04-03 07:22:56] Checking for reference FASTA file
[2017-04-03 07:22:56] Generating SAM header for test_ref
[2017-04-03 07:22:56] 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)
[2017-04-03 07:22:56] Mapping left_kept_reads to genome test_ref with Bowtie2 
[2017-04-03 07:22:56] Mapping left_kept_reads_seg1 to genome test_ref with Bowtie2 (1/3)
[2017-04-03 07:22:56] Mapping left_kept_reads_seg2 to genome test_ref with Bowtie2 (2/3)
[2017-04-03 07:22:56] Mapping left_kept_reads_seg3 to genome test_ref with Bowtie2 (3/3)
[2017-04-03 07:22:56] Mapping right_kept_reads to genome test_ref with Bowtie2 
[2017-04-03 07:22:56] Mapping right_kept_reads_seg1 to genome test_ref with Bowtie2 (1/3)
[2017-04-03 07:22:56] Mapping right_kept_reads_seg2 to genome test_ref with Bowtie2 (2/3)
[2017-04-03 07:22:57] Mapping right_kept_reads_seg3 to genome test_ref with Bowtie2 (3/3)
[2017-04-03 07:22:57] Searching for junctions via segment mapping
[2017-04-03 07:22:57] Retrieving sequences for splices
[2017-04-03 07:22:57] Indexing splices
Building a SMALL index
[2017-04-03 07:22:57] Mapping left_kept_reads_seg1 to genome segment_juncs with Bowtie2 (1/3)
[2017-04-03 07:22:57] Mapping left_kept_reads_seg2 to genome segment_juncs with Bowtie2 (2/3)
[2017-04-03 07:22:57] Mapping left_kept_reads_seg3 to genome segment_juncs with Bowtie2 (3/3)
[2017-04-03 07:22:57] Joining segment hits
[2017-04-03 07:22:57] Mapping right_kept_reads_seg1 to genome segment_juncs with Bowtie2 (1/3)
[2017-04-03 07:22:57] Mapping right_kept_reads_seg2 to genome segment_juncs with Bowtie2 (2/3)
[2017-04-03 07:22:57] Mapping right_kept_reads_seg3 to genome segment_juncs with Bowtie2 (3/3)
[2017-04-03 07:22:57] Joining segment hits
[2017-04-03 07:22:57] Reporting output tracks
-----------------------------------------------
[2017-04-03 07:22:57] A summary of the alignment counts can be found in ./tophat_out/align_summary.txt
[2017-04-03 07:22:57] Run complete: 00:00:01 elapsed
channsoden commented 6 years ago

It looks like this issue was fixed by Bowtie 2.3.1?

For future: Try installing the latest version of Bowtie.

http://bowtie-bio.sourceforge.net/bowtie2/index.shtml

nieder commented 6 years ago

Tophat-2.1.2 tests now pass with bowtie-2.3.4.1. At this point I'm not sure why I was testing against bowtie-2.3.0 when 2.3.1 was released slightly a few days before my comment. But I do know to try the latest code/releases when debugging if possible.

channsoden commented 6 years ago

Yes, not trying to snub you nieder, sorry. Just making a note for future people like myself who are trying to troubleshoot this issue.