Issue with alignement using Tophat2/Bowtie2 on MiRNA deep sequencing

Question

[cyrille.laurent.sage]

HI

I am trying to use Tophat2/Bowtie2 to align MiRNA from NGS to whole mouse genome. I am using tophat 2.1 and bowtie 2.1 . My command line is

bsub \
  -q priority \
  -W 240:00 \
  -n 8 \
    tophat2 \
    -G /Path/Reference-genes/genes.gtf \
    -p 8 \
    -N 7 \
    --read-edit-dist 7 \
    --library-type fr-unstranded \
    -o /Path/151013\ LIB18009\ seq/MiRNA-Ampli-test-101315D_output  \
    /Path/NGS_fq/Reference-genes/genome \
    /Path/151013\ LIB18009\ seq/MiRNA-Ampli-test-101315.fq

I am using a local server to run the job, but I got this answer back.

-----------------------------------------------
[2015-10-15 09:32:10] Checking for Bowtie
                                  Bowtie version:              2.1.0.0
[2015-10-15 09:32:10] Checking for Bowtie index files (genome)..
[2015-10-15 09:32:10] Checking for reference FASTA file
[2015-10-15 09:32:10] Generating SAM header for /groups/corey/Cyrille/NGS_fq/Reference-genes/genome
[2015-10-15 09:32:12] Reading known junctions from GTF file
[2015-10-15 09:32:16] Preparing reads
                 left reads: min. length=20, max. length=25, 920982 kept reads (46944 discarded)
[2015-10-15 09:32:25] Building transcriptome data files /groups/corey/Cyrille/151013 LIB18009 seq/MiRNA-Ampli-test-101315D_output/tmp/genes
[2015-10-15 09:32:38] Building Bowtie index from genes.fa
[2015-10-15 09:39:01] Mapping left_kept_reads to transcriptome genes with Bowtie2
                [FAILED]
Error running bowtie:
Warning: Output file 'seq/MiRNA-Ampli-test-101315D_output/tmp/genes' was specified without -S.  This will not work in future Bowtie 2 versions.  Please use -S instead.
Extra parameter(s) specified: "-"
Error: Encountered internal Bowtie 2 exception (#1)
Command: /opt/bowtie2-2.1.0/bowtie2-align --wrapper basic-0 -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,-44,0 -p 8 --sam-no-hd -x /Path/151013 LIB18009 seq/MiRNA-Ampli-test-101315D_output/tmp/genes - bowtie2-align exited with value 1

The issue is I do not know where to put -S option. If I put it at the beginning of the command line I got an error -S is not a Tophat2 option. If I tried the --b2-S it is not working. I am kind of stuck here

Thanks

Answer 1

[Ram]

I'd suggest removing the blank spaces in directory names, or using quotes around the directory name. Escape characters may be parsed but not passed along downstream by the shell.