Hello Yan,

Unfortunately, the tool can only extract sequence that is provided as the mapping target. This will be a problem with any of the methods. This tool does avoid a problem with generating negative coordinates (which will cause a problem with the 'Extract' tool). But it is not quite giving you what you want either, assuming that partially extended sequence, based on available data, would be acceptable.


Using the compute tool may be the best option for your case, now that the data is clearer. "End" coordinates that extend past the edge of the chromosome are not a problem, but the "Start" coordinate will need to be set to 1 (if using GFF3 as interval directly) or 0 (if you converted to BED - this doesn't appear to be the case). The expression below will either subtract '5000' from a "Start" coordinate or change it to a "1", depending on how close it is to the leading edge of the scaffold. (Modify for BED to be 0-based as needed).

(c2 - 5000) if (c2 > 5000) else (1)

Then add 5000 to the end, 'Cut' columns, and extract as Graham recommended.

I am not going to address the GFF3 format except to say that if you have gene rows in your data, use those if your target genome has spliced transcripts. If the data is transcript, not gene based, and is split between rows (multi-exon), then the processing becomes more complicated. One potential solution is the 'Extract' tool - it does not only extract fasta sequence, it can also be used to combine records for some GFF/GTF datasets - so you could try this and output "Interval" data instead of "Fasta". This creates a new GTF file with global coordinates (but the sequence output will be spliced). Check to see if correct, run the 'Compute' tool to do the extensions, 'Cut' columns, and do a final 'Extract' run to obtain the extended, global, sequence. All of this would have to be tested with your data - much depends on the attributes in your file.

Hopefully one of these solution will work out for you,

Jen
Galaxy team

On 9/25/12 12:41 AM, Yan He wrote:
Hi Jen,

Thanks very much for your help! It is very helpful. However, following
your suggestion, what I got is not what I want. Take one sequence for
example. The annotation for one scaffold is
C16582  GLEAN   mRNA    35      385     0.555898        -       .       
ID=OYG_GLEAN_10000001;
C16582  GLEAN   CDS     35      385     .       -       0       
Parent=OYG_GLEAN_10000001;

What I got for this scaffold is
 >?_C16582_385_5385_-
GCAAACAAGC
 >?_C16582_385_5385_-
GCAAACAAGC

                                                                
I understand that it is trying to get the sequence of the gene
downstream from 385-5385, but the sequence is short, so I only get what
the scaffold has. I would like to have the upstream+gene+downstream
sequence at the same time, not only the upstream or downstream. How can
I do this using a galaxy tool? Thanks!

Yan
                                                                



 > Date: Mon, 24 Sep 2012 12:26:03 -0700
 > From: j...@bx.psu.edu
 > To: yanh...@hotmail.com
 > CC: galaxy-user@lists.bx.psu.edu
 > Subject: Re: [galaxy-user] extract genome sequence
 >
 > Hi Yan,
 >
 > Both of the other suggestions are good - I'll also give you another
 > choice to build coordinates before using the "Fetch Sequences -> Extract
 > Genomic DNA" tool to obtain the fasta sequence.
 >
 > Using your input in BED/Interval format (convert from GFF/GTF if
 > necessary, using the tool "Convert Formats -> GFF-to-BED "), or the
 > first 6 columns if a BED12 (use "Cut" as needed), then run the "Operate
 > on Genomic Intervals -> Get flanks" tool.
 >
 > "Region:" Whole feature
 > "Location of the flanking region/s:" Both
 > "Offset" 0
 > "Length of the flanking region(s):" 5000
 >
 > Your question is similar to this one (the first part, but I thought you
 > might be interested in how to just get the flanks, too).
 >
http://user.list.galaxyproject.org/Get-flanks-version-1-0-0-td4604849.html
 >
 > Good luck with your project!
 >
 > Jen
 > Galaxy team
 >
 > ps. To search prior questions, please see:
 > http://galaxy.psu.edu/search/mailinglists/
 >
 > On 9/23/12 7:00 PM, Yan He wrote:
 > > Hi everyone,
 > >
 > > I have the genome sequence and gene annotation file. Is there a tool on
 > > Galaxy to extract the 5,000 bp upstream, 5,000 bp downstream and genome
 > > sequences of the genes (including exons and introns) from the genome
 > > sequence? Any suggestions are highly appreciated! Thanks!
 > >
 > > Yan
 > >
 > >
 > >
 > > ___________________________________________________________
 > > 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 Jackson
 > http://galaxyproject.org

--
Jennifer Jackson
http://galaxyproject.org
___________________________________________________________
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