Hi
Lately, I have a problem in running tophat 1.3.1 with a 100bp paired-end Illumina HiSeq RNA reads. After cleaning (quality trim, duplicate removal, adapter removal) I did split the files (taking care not to split the last entry sequence and quality scores) and fed to tophat . Please note, here I have more of left-kept reads because I have an extra file with leftover unpaired reads. Also, I have noticed with previous successful runs, eventhough the fed fastq paired read files have the same number of sequences what we see (in the log) as left-reads and right reads are slightly different.
Here is the log:
[Thu Oct 27 18:33:40 2011] Beginning TopHat run (v1.3.1)
-----------------------------------------------
[Thu Oct 27 18:33:40 2011] Preparing output location ./tophat_out/
[Thu Oct 27 18:33:40 2011] Checking for Bowtie index files
[Thu Oct 27 18:33:40 2011] Checking for reference FASTA file
[Thu Oct 27 18:33:40 2011] Checking for Bowtie
Bowtie version: 0.12.7.0
[Thu Oct 27 18:33:40 2011] Checking for Samtools
Samtools Version: 0.1.12a
[Thu Oct 27 18:33:40 2011] Generating SAM header for ../PG210SC5
[Thu Oct 27 18:33:40 2011] Preparing reads
format: fastq
quality scale: phred33 (default)
Left reads: min. length=50, count=134790672
Right reads: min. length=50, count=118121205
[Thu Oct 27 20:34:22 2011] Mapping left_kept_reads against PG210SC5 with Bowtie
[Thu Oct 27 21:42:15 2011] Processing bowtie hits
[Thu Oct 27 23:08:30 2011] Mapping left_kept_reads_seg1 against PG210SC5 with Bowtie (1/4)
[Fri Oct 28 00:27:19 2011] Mapping left_kept_reads_seg2 against PG210SC5 with Bowtie (2/4)
[Fri Oct 28 01:47:04 2011] Mapping left_kept_reads_seg3 against PG210SC5 with Bowtie (3/4)
[Fri Oct 28 02:57:47 2011] Mapping left_kept_reads_seg4 against PG210SC5 with Bowtie (4/4)
[Fri Oct 28 04:25:49 2011] Mapping right_kept_reads against PG210SC5 with Bowtie
[Fri Oct 28 05:26:52 2011] Processing bowtie hits
[Fri Oct 28 06:48:08 2011] Mapping right_kept_reads_seg1 against PG210SC5 with Bowtie (1/4)
[Fri Oct 28 08:00:12 2011] Mapping right_kept_reads_seg2 against PG210SC5 with Bowtie (2/4)
[Fri Oct 28 09:11:43 2011] Mapping right_kept_reads_seg3 against PG210SC5 with Bowtie (3/4)
[Fri Oct 28 10:21:22 2011] Mapping right_kept_reads_seg4 against PG210SC5 with Bowtie (4/4)
[Fri Oct 28 11:56:21 2011] Searching for junctions via segment mapping
[FAILED]
Error: segment-based junction search failed with err =1
____________________________________________________________________________________________
In the segment_juncs.log the last entry reads:
FZStream::rewind() popen(gzip -cd './tophat_out/tmp/left_kept_reads_seg1_missing.fq.z') failed
I have previously used such mixture of paired and unpaired reads successfully (I think!) with another set of reads. However, they were smaller read sets. Even with the above when I use only one pair out of four split files it works fine.
Appreciate if anyone can help me to resolve this problem.
Lately, I have a problem in running tophat 1.3.1 with a 100bp paired-end Illumina HiSeq RNA reads. After cleaning (quality trim, duplicate removal, adapter removal) I did split the files (taking care not to split the last entry sequence and quality scores) and fed to tophat . Please note, here I have more of left-kept reads because I have an extra file with leftover unpaired reads. Also, I have noticed with previous successful runs, eventhough the fed fastq paired read files have the same number of sequences what we see (in the log) as left-reads and right reads are slightly different.
Here is the log:
[Thu Oct 27 18:33:40 2011] Beginning TopHat run (v1.3.1)
-----------------------------------------------
[Thu Oct 27 18:33:40 2011] Preparing output location ./tophat_out/
[Thu Oct 27 18:33:40 2011] Checking for Bowtie index files
[Thu Oct 27 18:33:40 2011] Checking for reference FASTA file
[Thu Oct 27 18:33:40 2011] Checking for Bowtie
Bowtie version: 0.12.7.0
[Thu Oct 27 18:33:40 2011] Checking for Samtools
Samtools Version: 0.1.12a
[Thu Oct 27 18:33:40 2011] Generating SAM header for ../PG210SC5
[Thu Oct 27 18:33:40 2011] Preparing reads
format: fastq
quality scale: phred33 (default)
Left reads: min. length=50, count=134790672
Right reads: min. length=50, count=118121205
[Thu Oct 27 20:34:22 2011] Mapping left_kept_reads against PG210SC5 with Bowtie
[Thu Oct 27 21:42:15 2011] Processing bowtie hits
[Thu Oct 27 23:08:30 2011] Mapping left_kept_reads_seg1 against PG210SC5 with Bowtie (1/4)
[Fri Oct 28 00:27:19 2011] Mapping left_kept_reads_seg2 against PG210SC5 with Bowtie (2/4)
[Fri Oct 28 01:47:04 2011] Mapping left_kept_reads_seg3 against PG210SC5 with Bowtie (3/4)
[Fri Oct 28 02:57:47 2011] Mapping left_kept_reads_seg4 against PG210SC5 with Bowtie (4/4)
[Fri Oct 28 04:25:49 2011] Mapping right_kept_reads against PG210SC5 with Bowtie
[Fri Oct 28 05:26:52 2011] Processing bowtie hits
[Fri Oct 28 06:48:08 2011] Mapping right_kept_reads_seg1 against PG210SC5 with Bowtie (1/4)
[Fri Oct 28 08:00:12 2011] Mapping right_kept_reads_seg2 against PG210SC5 with Bowtie (2/4)
[Fri Oct 28 09:11:43 2011] Mapping right_kept_reads_seg3 against PG210SC5 with Bowtie (3/4)
[Fri Oct 28 10:21:22 2011] Mapping right_kept_reads_seg4 against PG210SC5 with Bowtie (4/4)
[Fri Oct 28 11:56:21 2011] Searching for junctions via segment mapping
[FAILED]
Error: segment-based junction search failed with err =1
____________________________________________________________________________________________
In the segment_juncs.log the last entry reads:
FZStream::rewind() popen(gzip -cd './tophat_out/tmp/left_kept_reads_seg1_missing.fq.z') failed
I have previously used such mixture of paired and unpaired reads successfully (I think!) with another set of reads. However, they were smaller read sets. Even with the above when I use only one pair out of four split files it works fine.
Appreciate if anyone can help me to resolve this problem.
Comment