Hello Lizex,

The utility tools from Tophat are not included with Galaxy, so that is perhaps why you are having trouble creating junctions files, but I am wondering if you need them at all.


You mention that you have different sequence types from different samples. These represent different conditions? Or some do and others are replicates or complete replacements for the originals (the re-sequenced data)?

For data representing different samples/conditions, you would not want to map the data together with Tophat or run it together in Cufflinks. Even replicates are not mapped in the same Tophat job, although they are included in the same Cufflinks job. The tool authors have an opinion about the value of replicates - so be sure to read about that at the Cufflinks web site.
http://cufflinks.cbcb.umd.edu/howitworks.html#reps

The first time different conditions would be in the same job would be at the stage where Cuffmerge is run, to prepare for Cuffdiff - where the differential expression analysis would take place. This is past the stage where individual reads are involved.

Our RNA-seq tutorial is here:
https://main.g2.bx.psu.edu/u/jeremy/p/galaxy-rna-seq-analysis-exercise

And several donated by the Galaxy community are here:
http://wiki.galaxyproject.org/Learn#Other_Tutorials

But the best resource is the paper from the tool authors ("Protocol"):
http://tophat.cbcb.umd.edu/manual.html
http://cufflinks.cbcb.umd.edu/manual.html

In the end, you may decide that some of the complexity of your data can be reduced by dropping some datasets, to simplify and achieve the same overall results, or perhaps even improved results. In general, I think it is probably safe to say that the less "done" to prep expression data, and certainly the more homogeneous it is, the better the result. But this is your decision.

Good luck with your project. Next time when asking a question, please open a brand new email message and start a new thread, not reply to an existing thread and just change the subject line. This helps us with tracking and is appreciated.

Thanks!

Jen
Galaxy team

On 2/16/13 5:23 AM, Lizex Husselmann wrote:
Dear all

I have in my project single end reads (50 bp) for some samples and
paired end reads (100 bp frw and rev) for  the other samples. I had to
re-sequence some of the samples of which I have paired-end reads.
However the re-sequence data I receives is single-end reads of 250 bp.
Tophat wont allow mapping single and paired-end reads together. It says
the result will look bad. They do mention that you can convert the
junctions.bed file (output of Tophat) with bed_to_juncs using the -j
option. I quote for TopHat manual "run TopHat on the 2nd set of reads
using the -j option to supply the junctions file produced by be_to_juncs
in the previous step". This is supposed to work but I didn't get it to
work. Any suggestion on how I should go about analyzing this data?

Kind regards

Lizex

 >>> Nate Coraor <n...@bx.psu.edu> 02/15/13 5:48 PM >>>
On Feb 15, 2013, at 10:35 AM, Mike Dufault wrote:

 > To whom it may concern:
 >
 > The History panel on the right side of the Galaxy page is taking a
very very long time to load. Also, when it does load, I have tired to
save my .bam files and the transmissions gets truncated to ~7000kb -
8000kb of data. All of my .bam files are several GB.
 >
 > Some times, when I retry tor download the data, it succeeds and other
times it is again truncated. The size of the truncation may be different
for the same file on the retry attempt.
 >
 > Is there a problem with Galaxy?

Hi Mike,

There are some performance problems with the Main site that we are
currently investigating Thanks for the information and we apologize for
the problems.

--nate

 >
 > Thanks,
 > Mike
 > ___________________________________________________________
 > 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:
 >
 > http://lists.bx.psu.edu/listinfo/galaxy-dev
 >
 > To manage your subscriptions to this and other Galaxy lists,
 > please use the interface at:
 >
 > http://lists.bx.psu.edu/


___________________________________________________________
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:

http://lists.bx.psu.edu/listinfo/galaxy-dev

To manage your subscriptions to this and other Galaxy lists,
please use the interface at:

http://lists.bx.psu.edu/

This message is confidential and may be covered by legal professional
privilege. It must not be read, copied, disclosed or used in any other
manner by any person other than the addressee(s). Unauthorised use,
disclosure or copying is strictly prohibited and may be unlawful. The
views expressed in this email are those of the sender, unless otherwise
stated. If you have received this email in error, please contact ARC
Service Desk immediately. (mailto:serviced...@arc.agric.za) To report
incidents of fraud and / or corruption in the ARC use our Ethics Hotline
by: Phone number : 0800 000 604 Fax number : 0800 00 7788 Email address
: a...@tip-offs.com
Please Call me : 32840
Website: www.tip-offs.com For more information on the ARC Ethics
Hotline, please visit our website at www.arc.agric.za.


___________________________________________________________
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:

   http://lists.bx.psu.edu/listinfo/galaxy-dev

To manage your subscriptions to this and other Galaxy lists,
please use the interface at:

   http://lists.bx.psu.edu/


--
Jennifer Hillman-Jackson
Galaxy Support and Training
http://galaxyproject.org
___________________________________________________________
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:

 http://lists.bx.psu.edu/listinfo/galaxy-dev

To manage your subscriptions to this and other Galaxy lists,
please use the interface at:

 http://lists.bx.psu.edu/

Reply via email to