Hello Carlos, Thanks a lot for the tip. The tab trick has fixed the problem. Regards, Aarti -----Original Message----- From: Carlos Borroto [mailto:carlos.borroto@gmail.com] Sent: Thursday, July 05, 2012 9:12 PM To: Avik Datta Cc: Aarti Desai; galaxy-user@lists.bx.psu.edu Subject: Re: [galaxy-user] Getting reference index files in local galaxy install Also make sure you are using TABs to separate the fields in the .loc file, this has bitten me several time in the past. My vim config places 4 spaces instead of TAB, to deactivate this option you can do ":set noexpandtab". Hope it helps, Carlos On Thu, Jul 5, 2012 at 4:39 AM, Avik Datta <reach4avik@gmail.com> wrote:

Hi Aarti,

Check the name of your ref file. If it is hg19.fa, then modify loc file as "hg19 hg19 HG19_BWA /root/Ref_INDEX/HG19BWAIndex/base/hg19.fa"

Avik Datta

On Thu, Jul 5, 2012 at 1:42 PM, Aarti Desai <aarti_desai@persistent.co.in> wrote:

...

Hi,

We have a local install of galaxy and I’m trying to add the reference index files for bwa using the information provided in the following link

http://wiki.g2.bx.psu.edu/Admin/NGS%20Local%20Setup

I have modified the bwa_index.loc file present in the ../tool-data directory by adding the path to where the index is on our server (Also attached). However, even after restarting the server, the reference genome does not show when choosing the “use a built-in index option”. I’m not sure whether the loc file is correctly created and whether any other configuration file needs to be changed/updated. Help in the matter greatly appreciated.

Thanks,

Aarti

From: galaxy-user-bounces@lists.bx.psu.edu [mailto:galaxy-user-bounces@lists.bx.psu.edu] On Behalf Of Jennifer Jackson Sent: Thursday, July 05, 2012 1:23 AM To: Lindsey Kelly Cc: galaxy-user@lists.bx.psu.edu Subject: Re: [galaxy-user] Initial QC and grooming for Illumina HiSeq2000 paired end RNAseq data

Hello Lindsey,

Yes, you have this correct. The general path would be to:

- join forward and reverse data per run - run FASTQ Groomer & FastQC (note: if your data is already in Sanger FASTQ format with Phred+33 quality scaled values, the datatype '.fastqsanger' can be directly assigned and the FASTQ Groomer step skipped. This is likely true if your data is a from the latest CASAVA pipeline, but please double check.) - discard data as needed based on quality - split forward and reverse data that passes QC - concatenate all forward reads from a sample into one FASTQ file - concatenate all reverse reads from a sample into one FASTQ file. - for each sample, run TopHat using the two concatenated FASTQ files

To manipulate paired end data, please see the tools -> NGS: QC and manipulation: FASTQ splitter & FASTQ joiner.

To combined data files head-to-tail from multiple runs into a single FASTQ file please see the tool -> Text Manipulation: Concatenate datasets.

I am not sure of the actual volume of data, but if these start to get large or TopHat errors with a memory problem, a local or cluster instance would be the recommendation: http://getgalaxy.org

For reference: http://tophat.cbcb.umd.edu/manual.html http://www.nature.com/nprot/journal/v7/n3/full/nprot.2012.016.html

Hopefully this helps. Others are welcome to post comments/suggestions.

Jen Galaxy team

On 7/2/12 11:17 AM, Lindsey Kelly wrote:

I am trying to do RNAseq analysis on Paired end data from the Hiseq2000. I have about 50 files for each sample (25 forward and 25 reverse - although each sample has a different number of files).

I think that I need to:

-convert them into FASTQ sanger format using the FASTSQ groomer tool

-check the quality using the FASTQqc tool

I don't know how to handle this many files. Do I have to groom and run the QC for each file? Should I join the paired files and run both tools on each pair, or should I combine all of the data for each sample (which I don't know how to do) and then groom and run the QC for all of the reads for the sample.

Thanks in advance for advice

Lindsey

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

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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:

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/

DISCLAIMER ========== This e-mail may contain privileged and confidential information which is the property of Persistent Systems Ltd. It is intended only for the use of the individual or entity to which it is addressed. If you are not the intended recipient, you are not authorized to read, retain, copy, print, distribute or use this message. If you have received this communication in error, please notify the sender and delete all copies of this message. Persistent Systems Ltd. does not accept any liability for virus infected mails.