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.gtf file issue - Error at parsing .tlst line (invalid strand):
I'm trying to align my reads to a genome + transcriptome, which is in the form of a .gtf file. It gets half way through and then runs this error:
Code:user@it053392:~$ export PATH=/home/user/Downloads/bowtie2:$PATH user@it053392:~$ export PATH=/home/user/Downloads/tophat2:$PATH user@it053392:~$ echo $PATH /home/user/Downloads/tophat2:/home/user/Downloads/bowtie2:/usr/local/sbin:/usr/local/bin:/usr/sbin:/usr/bin:/sbin:/bin:/usr/games:/usr/local/games user@it053392:~$ /home/user/Downloads/tophat2/tophat -G /home/user/Desktop/_sam/RNAseq_beta_data/Transcriptome/merged.remDup.gtf -o /home/user/Desktop/Tophat_out /home/user/Downloads/bowtie2/example/index/DinoAnt /home/user/Desktop/_sam/RNAseq_beta_data/trimmed/23Y70_trimmed.fastq [2015-11-04 11:52:45] Beginning TopHat run (v2.1.0) ----------------------------------------------- [2015-11-04 11:52:45] Checking for Bowtie Bowtie version: 2.2.6.0 [2015-11-04 11:52:45] Checking for Bowtie index files (genome).. [2015-11-04 11:52:45] Checking for reference FASTA file Warning: Could not find FASTA file /home/user/Downloads/bowtie2/example/index/DinoAnt.fa [2015-11-04 11:52:45] Reconstituting reference FASTA file from Bowtie index Executing: /home/user/Downloads/bowtie2/bowtie2-inspect /home/user/Downloads/bowtie2/example/index/DinoAnt > /home/user/Desktop/Tophat_out/tmp/DinoAnt.fa [2015-11-04 11:52:59] Generating SAM header for /home/user/Downloads/bowtie2/example/index/DinoAnt [2015-11-04 11:53:01] Reading known junctions from GTF file [2015-11-04 11:53:04] Preparing reads left reads: min. length=85, max. length=85, 23624572 kept reads (13384 discarded) [2015-11-04 12:00:30] Building transcriptome data files /home/user/Desktop/Tophat_out/tmp/merged.remDup [2015-11-04 12:00:51] Building Bowtie index from merged.remDup.fa [2015-11-04 13:22:45] Mapping left_kept_reads to transcriptome merged.remDup with Bowtie2 [FAILED] Error running: /home/user/Downloads/tophat2/bam2fastx --all /home/user/Desktop/Tophat_out/tmp/left_kept_reads.bam|/home/user/Downloads/bowtie2/bowtie2 -k 60 -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 /home/user/Desktop/Tophat_out/tmp/merged.remDup -|/home/user/Downloads/tophat2/fix_map_ordering --bowtie2-min-score 15 --read-mismatches 2 --read-gap-length 2 --read-edit-dist 2 --read-realign-edit-dist 3 --sam-header /home/user/Desktop/Tophat_out/tmp/merged.remDup.bwt.samheader.sam - - /home/user/Desktop/Tophat_out/tmp/left_kept_reads.m2g_um.bam | /home/user/Downloads/tophat2/map2gtf --sam-header /home/user/Desktop/Tophat_out/tmp/DinoAnt_genome.bwt.samheader.sam /home/user/Desktop/Tophat_out/tmp/merged.remDup.fa.tlst - /home/user/Desktop/Tophat_out/tmp/left_kept_reads.m2g.bam > /home/user/Desktop/Tophat_out/logs/m2g_left_kept_reads.out user@it053392:~$
Code:user@it053392:~$ /home/user/Downloads/tophat2/bam2fastx --all /home/user/Desktop/Tophat_out/tmp/left_kept_reads.bam|/home/user/Downloads/bowtie2/bowtie2 -k 60 -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 /home/user/Desktop/Tophat_out/tmp/merged.remDup -|/home/user/Downloads/tophat2/fix_map_ordering --bowtie2-min-score 15 --read-mismatches 2 --read-gap-length 2 --read-edit-dist 2 --read-realign-edit-dist 3 --sam-header /home/user/Desktop/Tophat_out/tmp/merged.remDup.bwt.samheader.sam - - /home/user/Desktop/Tophat_out/tmp/left_kept_reads.m2g_um.bam | /home/user/Downloads/tophat2/map2gtf --sam-header /home/user/Desktop/Tophat_out/tmp/DinoAnt_genome.bwt.samheader.sam /home/user/Desktop/Tophat_out/tmp/merged.remDup.fa.tlst - /home/user/Desktop/Tophat_out/tmp/left_kept_reads.m2g.bam > /home/user/Desktop/Tophat_out/logs/m2g_left_kept_reads.out Error at parsing .tlst line (invalid strand): 31958 TCONS_00032473 scaffold40. 5-1634 (ERR): bowtie2-align exited with value 141
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