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Galaxy error: can't fine fasta file
by Kenneth R. Auerbach 17 Sep '12

17 Sep '12

Hello, I'm new to Galaxy. When I read in a fasta file to Galaxy and then try to use it (in a blast search) as the query sequence, I get the error message below, although the uploaded fasta file is present in the history. Can anyone tell me what the problem could be? Is there some other step I need to do? Thank you. Error that appears under "nucleotide query sequence": ----- "History does not include a dataset of the required format / build" -----

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Galaxy CloudMan - Nodes can't make their own qsub calls?
by greg 17 Sep '12

17 Sep '12

Hi guys, I created a new Galaxy instance web launcher (https://biocloudcentral.herokuapp.com/launch) and then I ssh'd into the master node. I'm trying to run a Perl script that makes several qsub calls to other perl scripts. Now the catch is that one of those perl scripts makes its own qsub calls. And I'm getting this error when it tries to do that: Unable to run job: denied: host "ip-10-29-176-111.ec2.internal" is no submit host. Somehow this works fine on other clusters I've run this code on. Any idea what could be going on? Do I need to make all of the nodes "submit hosts"? Thanks a bunch! -Greg

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Does Tophat output *.accepted hits file contain headers?
by Du, Jianguang 14 Sep '12

14 Sep '12

Dear All, I want to use the Tophat output files with ".accepted hits" to do analysis outside Galaxy. However, the program I am using requires the Tophat output to be indexed, sorted BAM files that contain headers. Do the Tophat ouputs with ".accepted hits" produced at Galaxy contain headers? Will the headers of BAM files generated by Tophat universally the same? Thanks, Jianguang

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problem uploading via FTP server
by L.M. Slot 14 Sep '12

14 Sep '12

Dear Galaxy, I am using Galaxy on my work for analysing a whole genome sequencing project. On the server I wanted to upload two BAM files (ALL and FL). With FileZilla I uploaded them on the FTP server because the files are around 20 GB. When I tried to upload them in Galaxy I can see them in the list from the FTP server and when I try to upload them they appear in the column on the right, but they stay gray and the comment 'job is waiting to run' stays present instead of 'job running' or 'completed'. Hopefully you can help me with this problem. With kind regards, Linda Slot Linda Slot, MSc AMC Amsterdam Department of Pathology, L2-115 Meibergdreef 9 1105 AZ Amsterdam The Netherlands tel.nr. 020-5665638 What to include in a question 1. Where you are using Galaxy: Main<http://wiki.g2.bx.psu.edu/Main>, other public, local, or cloud instance 2. End-user questions from Test<http://wiki.g2.bx.psu.edu/Test> are generally not sent/supported - Test is for breaking 3. If a local or cloud instance, the distribution or galaxy-central hg pull # 4. If on Main<http://wiki.g2.bx.psu.edu/Main>, date/time the initial and ru-run jobs were executed 5. If there is an example/issue, exact steps to reproduce 6. What troubleshooting steps (if a problem is being reported) you have tested out 7. If on Main<http://wiki.g2.bx.psu.edu/Main>, you may be asked for a shared history link. Use Options → Share or Publish, generate the link, and email it directly back off-list. Note the dataset #'s you have questions about. 8. IMPORTANT: Get the quickest answer for data questions by leaving all of the input and output datasets in the analysis thread in your shared history undeleted until we have written you back. Use Options → Show Deleted Datasets and click dataset links to undelete to recover datasets if necessary 9. Always reply-all unless sharing a private link ________________________________ AMC Disclaimer : http://www.amc.nl/disclaimer ________________________________

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none availability genome.fa files through mapping modules on local server
by Sandrine Imbeaud 14 Sep '12

14 Sep '12

Hello, I apologize for this probably very simple application. We have installed our own Galaxy server and started using in-house the NGS modules. However, during the mapping procedure using either BWA for illumina or BFAST tools, no reference genome index is available. To solve the problem, we have followed the tutorials and have uploaded the hg19.fa file and put it locally in the Galaxy-dist/database folder. We also have modified the *_index.loc files indicating the path to the file. We restarted the Galaxy server. However, still no reference is available through the NGS mapping modules. Is there anyone that may help use solving this probably simple problem? Kind regards / Sandrine

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No output produced.....
by Neil.Burdett＠csiro.au 14 Sep '12

14 Sep '12

Hi, I have my own image registration tool that I've created on my own local instance of galaxy. The method takes in two images (*.nii.gz) formats and registers them together, and produces one registered *.nii.gz file and a *.trsf matrix file. The first issue encountered was the method was expecting *.nii.gz files as inputs but was receiving *.dat files. I navigated around this problem as shown by the files below: - <<tool id="RegisterAliBabaAffine" name="RegisterAffine"> < <description>two images</description> < <command interpreter="bash">$__root_dir__/tools/registration/reg-wrapper.sh $moving $fixed $outputTRSF $outputImage</command> - < <inputs> < <param format="binary" name="moving" type="data" label="Moving Image" /> < <param format="binary" name="fixed" type="data" label="Fixed Image" /> < <param type="hidden" name="outputTRSF" value="output.trsf" label="trsf file" help="Output File must have .trsf extension" /> < <param type="hidden" name="outputImage" value="output.nii.gz" label="Image output file" help="Output Image File must have .nii.gz extension" /> </inputs> - < <outputs> < <data format="input" name="output_TRSF" from_work_dir="output.trsf" /> < <data format="input" name="output_Image" from_work_dir="output.nii.gz" /> </outputs> < <help>This tool uses Affine Registration to register two images.</help> </tool> #!/bin/bash MOVING=`mktemp --suffix .nii.gz` FIXED=`mktemp --suffix .nii.gz` cat $1 > $MOVING cat $2 > $FIXED /usr/local/MILXView.12.08.1/BashScripts/RegisterAliBabaAffine -m $MOVING -f $FIXED -t $3 -o $4 RC=$? if [[ $RC == 0 ]]; then OUTPUTTRSF=`mktemp --suffix .trsf` OUTPUTIMG=`mktemp --suffix .nii.gz` cat $OUTPUTTRSF > $3 cat $OUTPUTIMG > $4 rm $OUTPUTTRSF rm $OUTPUTIMG fi rm $MOVING rm $FIXED exit $RC This allows them to pass the *.nii.gz files that the registration method is expecting. Everything works fine and I can see output generated in the job_working_dir and the history turns green... galaxy@bmladmin-OptiPlex-745:~$ ls -lrt ~/galaxy-dist/database/job_working_directory/000/27/ total 2940 -rw------- 1 galaxy nogroup 0 Sep 13 10:15 tmpRfHsOP_stderr -rw-r--r-- 1 galaxy nogroup 241 Sep 13 10:35 output.trsf -rw------- 1 galaxy nogroup 80 Sep 13 10:35 tmplmK0V2_stdout -rw-r--r-- 1 galaxy nogroup 2998272 Sep 13 10:38 output.nii.gz However, the problem occurs when the files are copied from ~/galaxy-dist/database/job_working_directory/000/27/ to ~/galaxy-dist/database/files/000/. When this happens the files become size = 0. Any ideas? -rw-r--r-- 1 galaxy nogroup 0 Sep 13 09:36 /home/galaxy/galaxy-dist/database/files/000/dataset_40.dat -rw-r--r-- 1 galaxy nogroup 0 Sep 13 09:36 /home/galaxy/galaxy-dist/database/files/000/dataset_41.dat -rw-r--r-- 1 galaxy nogroup 0 Sep 13 10:38 /home/galaxy/galaxy-dist/database/files/000/dataset_43.dat -rw-r--r-- 1 galaxy nogroup 0 Sep 13 10:38 /home/galaxy/galaxy-dist/database/files/000/dataset_42.dat The output in galaxy.log indicates it is successful: /home/galaxy/galaxy-dist/tools/registration/reg-wrapper.sh /home/galaxy/galaxy-dist/database/files/000/dataset_23.dat /home/galaxy/galaxy-dist/database/files/000/dataset_20.dat output.trsf output.nii.gz galaxy.jobs DEBUG 2012-09-13 10:38:10,334 The tool did not define exit code or stdio handling; checking stderr for success galaxy.jobs DEBUG 2012-09-13 10:38:10,361 finish(): Moved /home/galaxy/galaxy-dist/database/job_working_directory/000/27/output.trsf to /home/galaxy/galaxy-dist/database/files/000/dataset_42.dat as directed by from_work_dir galaxy.jobs DEBUG 2012-09-13 10:38:10,380 finish(): Moved /home/galaxy/galaxy-dist/database/job_working_directory/000/27/output.nii.gz to /home/galaxy/galaxy-dist/database/files/000/dataset_43.dat as directed by from_work_dir galaxy.jobs DEBUG 2012-09-13 10:38:10,609 job 27 ended Is the issue copying *.nii.gz files and *.trsf file into *.dat files? Anyway around this? I've also modified ~/galaxy-dist/lib/galaxy/jobs/__init__.py (line 363) to change shutil.move To shutil.copy2 (same results) Also put in a different output path to copy to. But essentially we have files with size in ~/galaxy-dist/database/job_working_directory/000/id/, but they files are size 0 after the move into ~/galaxy-dist/database/files/000 Thanks Neil

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Counting RNA-seq reads per class.
by Mohammad Heydarian 14 Sep '12

14 Sep '12

Hi All, I have been trying to count the number of RNA-seq reads that fall into the various Cufflinks class codes ('=', 'j', 'u', 'x', etc...) and I am curious how others are determining how to count reads per class.. I tried first using the BedTools tool where you "count" the number of reads overlapping another set of intervals and later realized that each interval is extended1 kb up and downstream prior to the analysis (by default and not adjustable on Galaxy), so the number of reads that were "counted" for all of the classes was always much more than the amount of reads that I had for my Bam file. I then tried to isolate reads from each class into separate BAM files, using the BedTools "intersect" tool and there I consistently end up with significantly less reads than I have in my sample. I am very curious to find out how others are tackling this problem on Galaxy. Thanks for any input! Cheers, Mo Heydarian

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Re: [galaxy-user] How can I extract sequence information fromm cuffdiff files?
by Jennifer Jackson 13 Sep '12

13 Sep '12

Hi Humberto, Yes, my apologies, this should have been included in the original reply. The 'locus' field in the Cuffdiff files refers to a gene bound - not individual transcripts. To get to the transcripts, the inputs to Cuffdiff need to be accessed. If you used Cuffmerge, the "merged transcripts" GTF file would be the correct file to use as input to "Extract". If you used just Cuffcompare, use the "combined transcripts" GTF. To know which transcript was associated with which gene bound, compare the Cuffmerge merged transcripts GTF attributes (9th column: gene_id, tss_id, etc) with Cuffdiffs "gene_id", "tss_id" values - is also in the test_id column, depending on the file. The Cuffcompare GTF comparisons will be similar. You can gain access to the GTF attributes with the tool "Filter and Sort -> Filter GTF data by attribute values_list". Cut out the column of interest in the Cuffdiff file ("Text Manipulation -> Cut"), edit as desired, and use as a list filter. Or explore the other GFF filter options in the same tool group. Take care, Jen Galaxy team On 9/13/12 11:14 AM, Humberto Boncristiani wrote: > Hi > > Fetch sequence-extract genomic DNA do not accept cuffidif files. > Should I convert this file to some specific format? > > Thanks, > > Humberto. > > *Dr. Humberto Boncristiani* > National Research Council (NRC) Fellow > Adjunct Research Associate > Department of Biology > Univ. North Carolina at Greensboro > 312 Eberhart Bldg > Greensboro, NC 27403, USA. > Tel.:(1) 336-256-2591 > Fax: (1) 336-334-5839 > email: humbfb(a)gmail.com <mailto:humbfb@gmail.com> > > > > > On Sep 13, 2012, at 2:06 PM, Jennifer Jackson wrote: > >> Hello, >> >> By no annotation, do you mean species-specific annotation (GTF) was >> not used? And you want to compare to a protein database like Genbank >> NR or RefSeq? Then these are the instructions. Please let us know if >> you had something else in mind. >> >> The sequence extraction can be done on Galaxy Main (if that is where >> you are working), but the BLAST will need to be run on a local or >> cloud install. To get set up (instance and data), start here: >> http://getgalaxy.org >> http://usegalaxy.org/cloud >> >> The BLAST+ wrapper recently moved from the distribution to the Tool >> Shed, but there are installation tools integrated to help get this >> into your instance. See the latest News Brief for details (Sept 7, >> 2012) - these are also good to follow as you maintain your instance: >> http://wiki.g2.bx.psu.edu/News >> http://wiki.g2.bx.psu.edu/DevNewsBriefs/2012_09_07 >> >> Questions about local/cloud installs are best directed to the >> galaxy-dev(a)bx.psu.edu mailing list: >> http://wiki.g2.bx.psu.edu/Mailing%20Lists >> >> To extract the transcript sequences, use the tool 'Fetch Sequences -> >> Extract Genomic DNA'. This will accept a custom reference genome from >> the history, if you have been using one, by changing the option >> "Source for Genomic Data:" to "History". >> >> Hopefully this helps, >> >> Jen >> Galaxy team >> >> On 9/13/12 10:09 AM, Humberto Boncristiani wrote: >>> Hi. >>> >>> I got cuffdiff files with gene differential expression on it. I don't >>> have the annotation, therefore I need to extract the sequence >>> information from the genome coordinates and them blast them to identify >>> those. >>> How the easiest way to do it? >>> >>> Thanks. >>> >>> Humberto >>> >>> >>> >>> *Dr. Humberto Boncristiani* >>> National Research Council (NRC) Fellow >>> Adjunct Research Associate >>> Department of Biology >>> Univ. North Carolina at Greensboro >>> 312 Eberhart Bldg >>> Greensboro, NC 27403, USA. >>> Tel.:(1) 336-256-2591 >>> Fax: (1) 336-334-5839 >>> email: humbfb(a)gmail.com <mailto:humbfb@gmail.com> >>> >>> >>> >>> >>> >>> >>> ___________________________________________________________ >>> 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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How can I extract sequence information fromm cuffdiff files?
by Humberto Boncristiani 13 Sep '12

13 Sep '12

Hi. I got cuffdiff files with gene differential expression on it. I don't have the annotation, therefore I need to extract the sequence information from the genome coordinates and them blast them to identify those. How the easiest way to do it? Thanks. Humberto Dr. Humberto Boncristiani National Research Council (NRC) Fellow Adjunct Research Associate Department of Biology Univ. North Carolina at Greensboro 312 Eberhart Bldg Greensboro, NC 27403, USA. Tel.:(1) 336-256-2591 Fax: (1) 336-334-5839 email: humbfb(a)gmail.com

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Tophat results
by Xiefan Fang 13 Sep '12

13 Sep '12

Dear galaxy users, I aligned my RNA-seq data by using Tophat in galaxy. It generated some "Tophat deletions", "Tophat insertions" and "Tophat splice junctions" results. These are all BED files. Does anyone know how to use/analyze these kind of results? Also, I used illumina RNA-seq. Each biological sample has 36-48 million reads. The data for each sample were divided to 10-12 FASTQ files. When I did the "FASTQ Summary Statistics" and draw "boxplot" for each of the sub-file, the score value is about 9-10. Is it too low? Shall I combine the FASTQ files for each biological sample and do the statistics again? At last, does anyone know how to convert a long list of zebrafish genes (500-1000 genes) to human or mammalian orthologs? Thank you for your replies, Xiefan Fang University of Florida

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