TopHat: no GFF3_file.expr in the output

[joseph]

Can you please help me to figure out why the “GFF3_file.expr” was not generated without getting any error message?
I had someone made me a GFF3 file from the KnowGene table; the first lines look like this:


##gff-version 3
gi|89161185|ref|NC_000001.9|NC_000001 UCSC gene 1115 4121 0 + 0 ID=uc001aaa.2gene
gi|89161185|ref|NC_000001.9|NC_000001 UCSC mRNA 1115 4121 0 + 0 ID=uc001aaa.2mRNA;Parent=uc001aaa.2gene
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 1115 2090 0 + 0 ID=uc001aaa.2exon1;Parent=uc001aaa.2mRNA
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 2475 2584 0 + 0 ID=uc001aaa.2exon2;Parent=uc001aaa.2mRNA
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 3083 4121 0 + 0 ID=uc001aaa.2exon3;Parent=uc001aaa.2mRNA
gi|89161185|ref|NC_000001.9|NC_000001 UCSC gene 1115 4272 0 + 0 ID=uc009vip.1gene
gi|89161185|ref|NC_000001.9|NC_000001 UCSC mRNA 1115 4272 0 + 0 ID=uc009vip.1mRNA;Parent=uc009vip.1gene
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 1115 2090 0 + 0 ID=uc009vip.1exon1;Parent=uc009vip.1mRNA
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 2475 4272 0 + 0 ID=uc009vip.1exon2;Parent=uc009vip.1mRNA
gi|89161185|ref|NC_000001.9|NC_000001 UCSC gene 4268 14764 0 - 0 ID=uc001aab.2gene
gi|89161185|ref|NC_000001.9|NC_000001 UCSC mRNA 4268 14764 0 - 0 ID=uc001aab.2mRNA;Parent=uc001aab.2gene
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 4268 4692 0 - 0 ID=uc001aab.2exon1;Parent=uc001aab.2mRNA
gi|89161185|ref|NC_000001.9|NC_000001 UCSC exon 4832 4901 0 - 0 ID=uc001aab.2exon2;Parent=uc001aab.2mRNA


I ran TopHat with the following command:

tophat -r 200 /myDir/bowtie-0.9.9.3/indexes/h_sapiens_asm s_7_1.fastq s_7_2.fastq -G GFF3_knownGene

[Cole Trapnell]

Hi Joseph,

All of the options need to be specified before the index and reads in the command. If you run it like this:

tophat -G GFF3_knownGene -r 200 /myDir/bowtie-0.9.9.3/indexes/h_sapiens_asm s_7_1.fastq s_7_2.fastq

You should get a non-empty .expr file.

[joseph]

Error: Report generation failed

Hi Joseph,

All of the options need to be specified before the index and reads in the command. If you run it like this:

tophat -G GFF3_knownGene -r 200 /myDir/bowtie-0.9.9.3/indexes/h_sapiens_asm s_7_1.fastq s_7_2.fastq

You should get a non-empty .expr file.

Hi Cole
I still don’t see an .expr file.
Here is the content of tophat_out
ls
GFF3_knownGene.juncs junctions.bed left_kept_reads.fq.candidate_hits.sam right_kept_reads.fq
accepted_hits.sam knownGene.juncs logs right_kept_reads.fq.candidate_hits.sam
coverage.wig left_kept_reads.fq repeat_reads.fa tmp


tophat -G GFF3_knownGene -r 200 /MyDir/bowtie-0.9.9.3/indexes/h_sapiens_asm s_7test_1.fastq s_7test_2.fastq

[Wed Jun 3 09:30:42 2009] Beginning TopHat run (v1.0.8)
-----------------------------------------------
[Wed Jun 3 09:30:42 2009] Preparing output location ./tophat_out/
[Wed Jun 3 09:30:42 2009] Checking for Bowtie index files
[Wed Jun 3 09:30:42 2009] Checking for reference FASTA file
[Wed Jun 3 09:30:42 2009] Checking for Bowtie
Bowtie version: 0.9.9.3
[Wed Jun 3 09:30:42 2009] Checking reads
seed length: 36bp
format: fastq
quality scale: phred
[Wed Jun 3 09:30:42 2009] Reading known junctions from GFF file
Warning: TopHat did not find any junctions in GFF file
Splitting reads into 1 segments
[Wed Jun 3 09:31:01 2009] Mapping reads against h_sapiens_asm with Bowtie
Splitting reads into 1 segments
[Wed Jun 3 09:31:12 2009] Mapping reads against h_sapiens_asm with Bowtie
[Wed Jun 3 09:31:21 2009] Searching for junctions via coverage islands
[Wed Jun 3 09:34:58 2009] Searching for junctions via mate-pair closures
[Wed Jun 3 09:39:55 2009] Retrieving sequences for splices
[Wed Jun 3 09:42:17 2009] Indexing splices
[Wed Jun 3 09:42:17 2009] Mapping reads against segment_juncs with Bowtie
[Wed Jun 3 09:42:18 2009] Joining segment hits
[Wed Jun 3 09:42:18 2009] Mapping reads against segment_juncs with Bowtie
[Wed Jun 3 09:42:18 2009] Joining segment hits
[Wed Jun 3 09:42:18 2009] Reporting output tracks
[FAILED]
Error: Report generation failed
M1-80330:test jdhahbi$

[Cole Trapnell]

Can you please email me the logs directory?