Vasu, Please reply to the mailing list as emails to individual Galaxy developers often get lost, and there are others on the list that might be able to help you or benefit from this discussion. Now, to your question: you're using the wrong GFF filtering tool, which is an easy mistake to make as there are many of them. You want to use Filter and Sort --> GFF --> Filter GTF data by attribute values list. Using this tool, I was able to filter dataset 11--a GTF file produced by Cuffcompare--using a Cuffdiff isoform expression file (dataset 10) on transcript_id. I've shared the modified history with you. Best, J. On Apr 18, 2011, at 3:29 PM, vasu punj wrote:

Hi Jeremy,

I have been trying to use the tool mentioned in this message. I have two samples comparison 6 and 5 and has run Cufflink/ Cuffcompare/ Cuffdiff. I have filtred the files for c12 i.e for significant analysis and file is 10 uploaded as B_A Cuffdiff isoform expr filtered.txt I uploaded the second file 11 B_A_Homo_sapiens.GRCh37.60.clean.combined which is a combined GTF file generated by cuffcompare.

When I tried to run filter combined transcript file using: Combined GTF as Cufflink assembled transcripts (11) and Cuffcompare tracking file as Cuffdiff isoform exp filtered file using sample no as 2, it return an empty file (12) Than thinking that perhaps it may be tracking file which I may have to use instead of combined GTF. I used B-A combined tracking file in place of combined GTF file but it will pop up only in Cuffcompare tracking file It may not be right but I used File 13 as tracking file with combined GTF as as assembled transcript still it return empty out put I have also shared history with you.

Would you like to point me what is going on here? Thanks.

Vasu --- On Mon, 4/11/11, Jeremy Goecks <jeremy.goecks@emory.edu> wrote:

From: Jeremy Goecks <jeremy.goecks@emory.edu> Subject: Re: [galaxy-user] downstream analysis of cuffdiff out put To: "shamsher jagat" <kanwarjag@gmail.com> Cc: "galaxy-user" <galaxy-user@lists.bx.psu.edu> Date: Monday, April 11, 2011, 9:04 AM

...

...

wrote: Jagat, Just like any mRNA-seq experiment to achieve following objectives: 1. Reconstruct all transcripts of a particular gene and corresponding Cuffdiff significantly expressed transcripts as called by cuffdiff. 2. What are different isoforms 3. Location of splicing

From various output files which unique ID can be matched from one file say Cuffdiff.expr (transcript/ isoform/Splicing) to other file - transcript.gtf corresponding to each sample or combined GTF file. I've got a script that does this for the cuffdiff isoform expression testing file and a GTF file; I'll wrap it up and add it to Galaxy in the next couple weeks. It would probably be useful to have similar scripts for the other expression testing files as well. Also, it would be nice to be able to take the FPKM values generated by Cuffdiff and attach them to their respective

On Thu, Mar 10, 2011 at 7:55 AM, Jeremy Goecks <jeremy.goecks@emory.edu transcripts as attributes.

Hello all,

I've added a tool called 'Filter GTF file by attribute values list' to the galaxy-central code repository. This tool is available on our test server ( http://test.g2.bx.psu.edu/ ) at Filter and Sort --> GFF --> Filter GTF data by attribute values list and will be available on our main server in the next few weeks.

As expected, this tool filters a GTF file based on a list of attribute values--or filters using a tabular file where attribute values are first column, as is the case for Cuffdiff output files. Potential attributes that can be filtered on include transcript_id, gene_id, tss_id, and p_id; conveniently, these are the IDs that Cuffdiff uses in its output files.

Here's an example workflow:

(1) Run Cufflinks/compare/diff (2) Filter Cufflinks isoform differential expression file for transcripts that are differentially expressed; in other words, filter for c12=='yes' (2) Use 'Filter GTF data by attribute values list' to filter Cuffcompare combined transcripts using the filtered file from step (2) as the attribute values list and, voila, you have a GTF file of the differentially expressed transcripts that you can view in your favorite genome browser.

Hope this helps; feedback is always welcome.

Best, J.

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