[varshacp]

Hi

I tried to run tohat using different gff file and genome sequence file and getting the same error

tophat -p 2 -G ca.gff3 -o cp05_thout1 caref_chr_pltd_unplaced1 cp05_ctl1.fastq

[2014-05-05 10:39:33] Beginning TopHat run (v2.0.9)
-----------------------------------------------
[2014-05-05 10:39:33] Checking for Bowtie
Bowtie version: 2.1.0.0
[2014-05-05 10:39:33] Checking for Samtools
Samtools version: 0.1.19.0
[2014-05-05 10:39:33] Checking for Bowtie index files (genome)..
[2014-05-05 10:39:33] Checking for reference FASTA file
[2014-05-05 10:39:33] Generating SAM header for caref_chr_pltd_unplaced1
format: fastq
quality scale: phred33 (default)
[2014-05-05 10:39:34] Reading known junctions from GTF file
[2014-05-05 10:39:39] Preparing reads
left reads: min. length=12, max. length=347, 18834597 kept reads (58333 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
[2014-05-05 10:45:43] Building transcriptome data files..
[FAILED]
Error: gtf_to_fasta returned an error.

Thankx for your help

Varsha

[GenoMax]

HI

I checked the log file and besides the run.log which I posted earlier I get the following error in the g2f.log file

terminate called after throwing an instance of 'std:ut_of_range'
what(): basic_string::substr


Help me to understand this

Thank you
Varsha

The problem almost certainly seems to be the gff file (since all other index files appear to be there and I assume are non-zero byte).

Can you try to verify your GFF file using one of these: http://genometools.org/cgi-bin/gff3validator.cgi or http://modencode.oicr.on.ca/cgi-bin/...te_gff3_online

[jernest1]

Is the gtf_to_fasta executable in your PATH?
enter "echo $PATH" and make sure that gtf_to_fasta program (in whatever tophat folder was created when you unpacked the tar.gz file) is in one of those directories. $HOME/bin is a convenient directory for this.

Something else I have to do on my computer system (sun grid engine HPC cluster) is use qsub to submit jobs. The system has trouble finding all the software, so I submit jobs using "qsub -v PATH job.sge".

Maybe that will help someone.

[DRAT]

Hi hathiram2,

I recently had the same problem after downloading mm10 sequences/annotation files from the tophat website. Check the content of the genome.fa file because for me it was empty and I had to download it separately from the NCBI website. After replacing genome.fa file things work fine.