Hi Carlos, Thank you very much for this explanation. The format of my intervals file is: chr133289059732890664NM_000059_cds_1_0_chr13_32890598_f0+chr1332893213 32893462NM_000059_cds_2_0_chr13_32893214_f0+chr133289921232899321 NM_000059_cds_3_0_chr13_32899213_f0+chr133290023732900287 NM_000059_cds_4_0_chr13_32900238_f0+etc... Can you please explain me how to change this format so I will be able to give it as an input to DepthOfCoverage Thanks, Lilach 2012/6/21 Carlos Borroto <carlos.borroto@gmail.com>
Hi Jennifer, Thank you for this reply.
I made a new BWA file, this time using the hg19(full) genome. However, when I am trying to use DepthOfCoverage, the reference genomr is stucked on the hg_g1k_v37 (this is the only option to select), and I cannot change it to hg19(full). Most probably, because I selected hg_g1k_v37 in
On Thu, Jun 21, 2012 at 10:50 AM, Lilach Friedman <lilachfr@gmail.com> wrote: the
previous time I tried to use DepthOfCoverage. It seems as a bug? How can I change it?
Hi Lilach,
I have been dealing with these issues for some time now.
The only genome you can use with Picard and GATK tools in Galaxy is hg_g1k_v37. I think this is why.
From GATK Wiki[1]: "If you are using human data, your reads must be aligned to one of the official b3x (e.g. b36, b37) or hg1x (e.g. hg18, hg19) references. The contig ordering in the reference you used must exactly match that of one of the official references canonical orderings. These are defined by historical karotyping of largest to smallest chromosomes, followed by the X, Y, and MT. The order is thus 1, 2, 3, ..., 10, 11, 12, ... 20, 21, 22, X, Y, MT. The GATK will detect misordered contigs (for example, lexicographically sorted) and throw an error. This draconian approach, though unnecessary technically, ensures that all supplementary data provided with the GATK works correctly. You can use ReorderSam to fix a BAM file aligned to a missorted reference sequence."
[1] http://www.broadinstitute.org/gsa/wiki/index.php/Input_files_for_the_GATK
So far what I have done when presented with a BAM file produced with reference with lexicographical chromosomes ordering, is to use Picard's ReorderSam tool, also in Galaxy, selecting hg_g1k_v37 as reference. You might not be able to this, as if a recall correctly hg19 also use chr1, chr2... instead of 1, 2, ... In that case more work needs to be done and at that point is almost easier to just remap with the correct reference for use with GATK. In your case it seems you already have it. What you might need to do is resort your intervals file and probably change the chromosomes identifiers, this I think can be done inside Galaxy.
I would love to hear comments about this approach, as sometime I do worry like Hiram's comment hints to, that hg19 and hg_g1k_v37 might not be completely identical beside the chromosome ordering. In that case my resorted BAM or intervals files might be incorrect.
Hope it helps, Carlos
Thanks, Lilach
2012/6/18 Jennifer Jackson <jen@bx.psu.edu>
Hi Lilach,
The problem with this analysis probably has to do with a mismatch
the genomes: the intervals obtained from UCSC (hg19) and the BAM from your BWA (hg_g1k_v37) run.
UCSC does not contain the genome 'hg_g1k_v37' - the genome available from UCSC is 'hg19'.
Even though these are technically the same human release, on a practical level, they have a different arrangement for some of the chromosomes. You can compare NBCI GRCh37 with UCSC hg19 for an explanation. Reference genomes must be exact in order to be used with tools - base for base. When they are exact, the identifier will be exact between Galaxy and the
(UCSC, Ensembl) or the full Build name will provide enough information to make a connection to NCBI or other.
Sometimes genomes are similar enough that a dataset sourced from one can be used with another, if the database attribute is changed and the data from the regions that differ is removed. This may be possible in your case, only trying will let you know how difficult it actually is with your analysis. The GATK pipeline is very sensitive to exact inputs. You will need to be careful with genome database assignments, etc. Following the links on
between source the
tool forms to the GATK help pages can provide some more detail about expected inputs, if this is something that you are going to try.
Good luck with the re-run!
Jen Galaxy team
On 6/18/12 4:42 AM, Lilach Friedman wrote:
Hi, I am trying to used Depth of Coverage to see the coverages is specific intervals. The intervals were taken from UCSC (exons of 2 genes), loaded to Galaxy and the file type was changed to intervals.
I gave to Depth of Coverage two BAM files (resulted from BWA, selection of only raws with the Matching pattern: XT:A:U, and then SAM-to-BAM) and the intervals file (in advanced GATK options). The consensus genome is hg_g1k_v37.
I got the following error message:
An error occurred running this job: Picked up _JAVA_OPTIONS: -Djava.io.tmpdir=/space/g2main ##### ERROR
##### ERROR A USER ERROR has occurred (version 1.4-18-g80a4ce0): ##### ERROR The invalid argume
Is it a bug, or did I do anything wrong?
I will be grateful for any help.
Thanks! Lilach
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-- Jennifer Jackson http://galaxyproject.org
___________________________________________________________ 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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