Thanks Mete and Jenifer for your information.

Last time, I did mRNA sequencing analysis and decided to go with TopHat>Htseq>DESeq for DE genes. Just because the results match up quite nicely with my qPCR validation. Although Cuufflink/Cuffdiff produced results with quite similar trend with DESeq, It seem to me that Cuffdiff tend to inflate the fold-change and is not good at statistical analysis. Thanks Jenny and Ross.

Anyway, about the miRNA I am working on now. My miRNA data is 51bp, single-ended. I am going to cut adapter using FASTX-toolkit and align reads using Novoalign ( as Mete suggests) and Bowtie.

Just have a question for now: my 3' adapter sequence is : 5-rAppAGATCGGAAGAGCACACGTCT-NH2-3. How many bases should I put in the " Enter custom clipping sequence" ? Is " AGATCGGA" is optimal? Just because I observed that many reads only contain part of 3' adapter sequence.

Also, Do I need to trim 5' adapter as well? and How?

Thank so much for your help

Thanh

On Wed, Sep 18, 2013 at 8:12 PM, Jennifer Jackson <jen@bx.psu.edu> wrote:

Hi Thanh,

Did the Tuxedo suite not work out for you in the end? Or the other tools that Ross suggested? These are both pipelines that are in common use.
http://lists.bx.psu.edu/pipermail/galaxy-user/2013-July/006367.html

Using a cloud Galaxy and installing tools from the Tool Shed is required for certain tools, perhaps that is the problem? Many tools now have automatic dependency installation, making set-up much easier. For a demonstration, watch the Channel: Galaxy ToolShed videos at Vimeo:
http://vimeo.com/user20484153

You also may want to look at some of the miRNA specific tools in the Tool Shed. They can be found under "Sequence Analysis". Most of these have online documentation, or the tool author includes documentation, that you can review to see if the tool is a good fit for what you want to do (if it is not expression analysis anymore, or you want to try something different like DESeq).
http://toolshed.g2.bx.psu.edu/repository

Hopefully this helps,

Jen
Galaxy team

On 9/18/13 10:19 AM, Hoang, Thanh wrote:
Hi all,
I would like to analyze my miRNA sequencing analysis from mouse tissue. I have not any idea which tools or pipeline work best. Do you have any suggestion?

Regards

Thanh
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