Hi Giuseppe,
Your workflow sounds correct. Cuffmerge or Cuffcompare is needed and no,
you do not want to use Cufflinks as sample input - these should be
mapped reads as you state that you have used them so far.
Are you using the public Main Galaxy server at http://main.g2.bx.psu.edu
(usegalaxy.org)? If so and you would like to share a history, I can take
a quick look to see if any usage problems are present.
For the overall results - there are many factors that can contribute to
different results vs what is published.
It may be that parameters for some of these steps need to be tuned, or
that obtaining the same results without a reference annotation to guide
assembly is not possible. We can offer help torule out anything
technically wrong with the workflow, so you can concentrate on these
scientific questions (test/try alternate parameter settings and such).
The paper from the tool authors (in the link below) would be a good
place to start once you get to this point, if you are not already using it.
To share, use the gear icon at the top of the history panel (upper right
corner), select "Share or Publish", click on the first button to
generate the share link, copy and paste that into a reply email to just
me (not the entire list, to keep your data private).
If instead you want to just explore this more on your own, links to help
for RNA-seq tools are in this wiki:
http://wiki.galaxyproject.org/Support#Interpreting_scientific_results
See Tools on the Main server: Example ? RNA-seq analysis tools.
Best,
Jen
Galaxy team
On 4/16/13 7:38 AM, Ianiri, Giuseppe wrote:
Hi,
I want to compare gene deferentially expressed in two conditions (1
and 2). I don't have a GTF reference annotation.
In my first attempt, I mapped my Illumina reads with TopHat. Then I
run Cufflink, and Cuffmerge on my Cufflink outputs for the conditions
1 and 2. At the end, I run Cuffdiff using the Cuffmerge output (1 + 2)
and the TopHat output for the two samples of interest. I got genes and
transcripts differentially expressed, but I was expecting a quite
different output (based on the literature).
Do you guys think I did anything wrong?
Also, I am not sure the question makes sense, but is the Cuffmerge
step essential for the comparison of my two samples? Or could I use my
Cufflink outputs for samples 1 and 2 for running Cuffdiff?
Regards,
Giuseppe Ianiri
Email: iani...@umkc.edu
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___________________________________________________________
The Galaxy User list should be used for the discussion of
Galaxy analysis and other features on the public server
at usegalaxy.org. Please keep all replies on the list by
using "reply all" in your mail client. For discussion of
local Galaxy instances and the Galaxy source code, please
use the Galaxy Development list:
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please use the interface at:
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To search Galaxy mailing lists use the unified search at:
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