Tophat gtf problem

[Jetse]

Hello everyone,

I am using tophat with a gtf file. I used it once with this command:

/home/sge_share_fedor8/common_scripts/tophat-2.0.6/tophat2 --color --quals  --GTF /home/sge_share_fedor12/jetse/GENOMES/GRCh37_gatk/human_GRCh37_gatk.gtf -x 1 --library-type fr-secondstrand --read-realign-edit-dist 0 -N 10 -m 2 -a 5 -p 5 --read-edit-dist 10 --coverage-search --bowtie1 -o /home/sge_share_fedor12/jetse/wntSignalling/test//tophat/PolyPlus1/ /home/sge_share_fedor12/jetse/GENOMES/GRCh37_gatk/GRCh37_gatk_colorspace /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F3.csfasta /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F5.csfasta /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F3.qual /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F5.qual

After this I want to run Tophat 26 times, so I dont want to build this gtf file everytime again. For this I use the command:

/home/sge_share_fedor8/common_scripts/tophat-2.0.6/tophat2 --color --quals  --transcriptome-index /home/sge_share_fedor12/jetse/GENOMES/GRCh37_gatk/human_GRCh37_gatk -x 1 --library-type fr-secondstrand --read-realign-edit-dist 0 -N 10 -m 2 -a 5 -p 5 --read-edit-dist 10 --coverage-search --bowtie1 -o /home/sge_share_fedor12/jetse/wntSignalling/test//tophat/PolyPlus1/ /home/sge_share_fedor12/jetse/GENOMES/GRCh37_gatk/GRCh37_gatk_colorspace /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F3.csfasta /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F5.csfasta /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F3.qual /data/jetse/wntSignalling/rawData/csfasta/polyplus1/run1/reads/PolyPlus1_F5.qual

This is exactly the same command, but i removed the .gtf and changed the --GTF to --transcriptome-index. But now I get this error:

[2013-02-12 15:15:48] Beginning TopHat run (v2.0.6)
-----------------------------------------------
[2013-02-12 15:15:48] Checking for Bowtie
                  Bowtie version:        0.12.8.0
[2013-02-12 15:15:48] Checking for Samtools
                Samtools version:        0.1.14.0
Error: GFF transcripts file not found or invalid (/home/sge_share_fedor12/jetse/GENOMES/GRCh37_gatk/human_GRCh37_gatk.gff)

When I look at the directory where the indexes are build, all references are build when using the first command. When this command is finished, the indexes are removed! What am I doing wrong? I just want to build an index once of my gtf file and want to use the index after the first time...

[Jetse]

Had to read the manual better...

TopHat should be first run with the -G option and with the --transcriptome-index option

I hope this will fix it