Running tophat with unmated pairs

[thinkRNA]

There seems to be no description of how to run tophat if you are using unpaired sequence reads. Does it only work for mated pairs?

When I give it my commands, this is the error:
Error: you must set the mean inner distance between mates with -r

But, I don't have mates?

[simonandrews]

There seems to be no description of how to run tophat if you are using unpaired sequence reads. Does it only work for mated pairs?

No, you can definitely use it for single end sequences. You just supply one sequence file instead of two.

tophat /path/to/indices fastq_file

[Cole Trapnell]

As noted in the manual, multiple (single end) FASTQ files can be provided as a comma separated list. When you have paired end reads, you need *two* comma separated lists.

[thinkRNA]

Thanks, I did get it going. But now, I get this error:

[Thu Feb 11 09:37:18 2010] Preparing output location ./tophat_out/
[Thu Feb 11 09:37:18 2010] Checking for Bowtie index files
[Thu Feb 11 09:37:18 2010] Checking for reference FASTA file
[Thu Feb 11 09:37:18 2010] Checking for Bowtie
Bowtie version: 0.11.3.0
[Thu Feb 11 09:37:18 2010] Checking reads
seed length: 27bp
format: fastq
quality scale: phred33 (default)
[Thu Feb 11 09:37:18 2010] Mapping reads against mm9 with Bowtie
[Thu Feb 11 09:37:21 2010] Joining segment hits
[Thu Feb 11 09:37:21 2010] Searching for junctions via segment mapping
Warning: junction database is empty!
[Thu Feb 11 09:38:12 2010] Joining segment hits
[Thu Feb 11 09:38:13 2010] Reporting output tracks

Does any one know how to fix this warning? I am using UCSC mm9 and thought the chromosome naming problem should disappear.

[middlemale]

in my case, single end read and I try to use gene model annotation
tophat -G <GFF3 file> /path/to/indices fastq_file

same problem "Error: you must set the mean inner distance between mates with -r"

and if without -G <GFF3 file> in options, that works fine.

any suggest of how adding -G option? cheers

[Thomas Doktor]

middlemale, could you copy-paste the exact command you used to launch TopHat, as you've typed it?