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

 

 

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

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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