Hello Stanislas,

To use a custom reference genome with Bowtie2/Tophat2, the genome needs to be loaded using FTP into your history in .fasta format. UCSC is one source, in the downloads area. It will require a download from there first. http://genome.ucsc.edu or from our rsync server. Just be aware that rat may be too large to run as a custom genome on the public server with this tool (I haven't tried it). If the tool ends with a memory error, then that is the problem.

We do have plans to add more indexes genomes for Bowtie2/Tophat2, but the public server migration is the priority right now. There are alternate wrappers available for Tophat (v1) in the Tool Shed (for SOLiD), and also Tophat2, for use in a local, cloud, or slipstream Galaxy. Our test server (https://test.galaxyproject.org/) has Tophat for SOLiD loaded, but this server is where we really do test, and quotas are small (10G) - so, tool problems are not supported and it is not recommended unless for test/trial runs. Both tools have links to documentation, with the Tophat2 website being a great resource to better understand how the tools differ (along with the google support group).

I would estimate that more indexes will be available starting in about a month, but it could be sooner/later.


Galaxy team

On 10/12/13 1:33 PM, Stanislas Werfel wrote:

I am new to the field of RNA-Seq analysis. I'd be very greatful if somene could help me with the following issue:

I'm using the Galaxy Main plattform and would like to run Tophat2 on RNA-Seq data from rat. However in Main under Tophat2 there are only built-in human and mouse genomes, while a lot more genomes are vailable under Tophat for Illumina.
I tried to import the rat genome sequence using the built-in UCSC Table browser, however I only managed to get a full genome as one FASTA file with all the different sequences. After mapping to this genome I can't see any hits when I click "display at UCSC main" on accepted hits. So I assume that the mapped reads do not conform to the official reference rat genome (rn5).

So how can one solve this isse?
-Is there a way to import a proper genome for use with Tophat2? Ideally directly indo Galaxy without prior download.
-If not, how does Tophat2 differ from "Tophat for Illumina". Can one also analyze non-Illumina data with that? As I understand it Tophat2 is faster than the previous version?

Thanks in advance for any help.

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Jennifer Hillman-Jackson