Re: [galaxy-user] Cuffdif output NOTEST
Hi Meike, Sorry for the delay. The Cuffdiff manual (http://cufflinks.cbcb.umd.edu) has help for interpreting NO TEST/LOW DATA results, and for adjusting the -c option - these are most often related to low coverage and/or fragmented transcripts. I am wondering, did you run Cufflinks to assemble the data first? This wiki section has help for recommended protocols (and includes links back to the tool's site above): https://wiki.galaxyproject.org/Support#Interpreting_scientific_results See - Tools on the Main server: RNA-seq As explained, it could be that there is a mismatch between the reference annotation file and your mapped data. The gtf file contains chromosome identifiers using Ensembl's nomenclature, while the built-in reference genome used for mapping rn5 is sourced from UCSC and uses their nomenclature. These are formatted differently. An exact match is required between identifiers or the annotation will be effectively be ignored. Often adding a chr to the start of the Ensembl chromosome name will resolve the match, but not always. The source for rn5 was here: http://hgdownload.cse.ucsc.edu/goldenPath/rn5/bigZips/ iGenomes is the preferred reference annotation source, due to the inclusion of all the attributes specifically used by the Cuffdiff tool and how these are created with specific data sources in mind (adjusted for their identifier nomenclature). This build is not yet available, but rn4 is: http://cufflinks.cbcb.umd.edu/igenomes.html Checking protocol and that the identifiers are a match are the first steps. Examine parameter tuning after. I didn't find this data in any of the histories you already shared, but the above help will resolve/explain most issues or results from this pipeline. Best, Jen Galaxy team On 4/8/14 5:46 AM, meike.l...@mdc-berlin.de wrote: Dear all, I have Illumina RNAseq data and want to look for differences in gene expression between male and female rats and transgenic vs. wildtype; for each condition I have triplicates. I mapped with TopHat for Illumina, using the reference genome rn5 and default settings. I did Cuffdiff afterwards and used the GTF-file Rattus norvegicus.Rnor_5.0.72.gtf as transcript. As result I got no significant changes in expression and it always says NO TEST (or sometimes LOW DATA). I found that lowering or raising the -c option might control the cuffdiff behavior, but I do not know what it is and how to control it. Can you explain me how to do it? And would you advise me to use another transcript for Cuffdiff? And when yes, where can I get it? Lots of questions. Thanks in advance for your answer. Best, Meike ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ -- Jennifer Hillman-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: 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
[galaxy-user] Base Recalibration tool on usegalaxy
Dear all, I can't find either of Table Recalibration or Base Recalibrator on usegalaxy public server. I am defident this is right place to ask, but seqanswers is clouded with many other pipelines. Is there an update of base recalibration tool on usegalaxy? Best, Kaz ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
Re: [galaxy-user] Base Recalibration tool on usegalaxy
Hi Kaz, you are searching for GATK tools, right? Please have a look at the GATK suites in the Tool Shed. Cheers, Bjoern Am 12.04.2014 00:16, schrieb KS: Dear all, I can't find either of Table Recalibration or Base Recalibrator on usegalaxy public server. I am defident this is right place to ask, but seqanswers is clouded with many other pipelines. Is there an update of base recalibration tool on usegalaxy? Best, Kaz ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
[galaxy-user] trouble visualizing data
Hi Lindsey, I see a few potential problem areas here. Double check these things: 1. when uploading, if the files are over 2G, or close to that, use FTP instead of a browser upload. Here is how: https://wiki.galaxyproject.org/FTPUpload 2. do not use the option to convert spaces to tabs. I doubt this does anything to a compressed format like BAM, but you almost never want to use this. Really is only for hand entered or pasted in data. 3. Try to convert the file to SAM to see if that functions. This will do two things: confirm basic BAM format is intact and permit you to examine the sequence identifiers on the header. Or try a tool like Picard's 'BAM Index Statistics'. 4. There could be an identifier mismatch problem between your data and our internal reference genome. The only ways to check are to either run some type of job that will list out our genome's sequences in the UI (like a small mapping job that creates a Galaxy-native BAM/SAM file) or obtain our version of the genome by rysnc and compare your files against it locally (before uploading to Galaxy). Both have advantages. The rsync server instructions are here, and I happened to use the genome question from yesterday to fill in the example last night, so you have near-exact instructions! https://wiki.galaxyproject.org/Admin/UseGalaxyRsync 5. If all else fails, consider loading the genome that you used to do the mapping up to Galaxy as a Custom reference genome: https://wiki.galaxyproject.org/Support#Custom_reference_genome https://wiki.galaxyproject.org/Support#Reference_genomes There have been some UI delays, but these are transient. Resubmitting the action has been working, so try that if buttons do not respond the first time. Let us know if you continue to have problems, Jen Galaxy team On 4/11/14 9:20 AM, Lindsey Fallis wrote: Hi Jennifer, I’m a post doc working with Brenda Oppert (she contacted you yesterday with some problems). I too have been having some problems getting visualizations to work. My goal is to show the Tribolium genome as a Circster plot with my RNA-seq data laid on top. My RNA-seq data is currently in .bam format. So far what I have a attempted to do is upload my .bam files into Galaxy using the Get Data, upload file from computer functions. Then I choose my file, check the ‘convert spaces to tabs’ box and set the genome to Tcas. It uploads correctly. Then I ask it to visualize as Circster and nothing happens… Then I tried viewing in trackster and nothing happens OR I get an error message saying one of my sequences isn’t in the genome file. Any suggestions to get the visualizations to work so that I have the Tcas chromosomes as the Circster backbone and my RNA-seq data showing around it? Thank you! Lindsey -- Jennifer Hillman-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: 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
[galaxy-user] trouble visualizing data
Hi Lindsey, I see a few potential problem areas here. Double check these things: 1. when uploading, if the files are over 2G, or close to that, use FTP instead of a browser upload. Here is how: https://wiki.galaxyproject.org/FTPUpload 2. do not use the option to convert spaces to tabs. I doubt this does anything to a compressed format like BAM, but you almost never want to use this. Really is only for hand entered or pasted in data. 3. Try to convert the file to SAM to see if that functions. This will do two things: confirm basic BAM format is intact and permit you to examine the sequence identifiers on the header. Or try a tool like Picard's 'BAM Index Statistics'. 4. There could be an identifier mismatch problem between your data and our internal reference genome. The only ways to check are to either run some type of job that will list out our genome's sequences in the UI (like a small mapping job that creates a Galaxy-native BAM/SAM file) or obtain our version of the genome by rysnc and compare your files against it locally (before uploading to Galaxy). Both have advantages. The rsync server instructions are here, and I happened to use the genome question from yesterday to fill in the example last night, so you have near-exact instructions! https://wiki.galaxyproject.org/Admin/UseGalaxyRsync 5. If all else fails, consider loading the genome that you used to do the mapping up to Galaxy as a Custom reference genome: https://wiki.galaxyproject.org/Support#Custom_reference_genome https://wiki.galaxyproject.org/Support#Reference_genomes There have been some UI delays, but these are transient. Resubmitting the action has been working, so try that if buttons do not respond the first time. Let us know if you continue to have problems, Jen Galaxy team On 4/11/14 9:20 AM, Lindsey Fallis wrote: Hi Jennifer, I’m a post doc working with Brenda Oppert (she contacted you yesterday with some problems). I too have been having some problems getting visualizations to work. My goal is to show the Tribolium genome as a Circster plot with my RNA-seq data laid on top. My RNA-seq data is currently in .bam format. So far what I have a attempted to do is upload my .bam files into Galaxy using the Get Data, upload file from computer functions. Then I choose my file, check the ‘convert spaces to tabs’ box and set the genome to Tcas. It uploads correctly. Then I ask it to visualize as Circster and nothing happens… Then I tried viewing in trackster and nothing happens OR I get an error message saying one of my sequences isn’t in the genome file. Any suggestions to get the visualizations to work so that I have the Tcas chromosomes as the Circster backbone and my RNA-seq data showing around it? Thank you! Lindsey -- Jennifer Hillman-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: 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
Re: [galaxy-user] Base Recalibration tool on usegalaxy
Hi Bjoern, Thank you for the reply. I found Table Recalibration, probably from GATK, on Tool Shed as you informed. http://toolshed.g2.bx.psu.edu/repository/view_repository?sort=nameadvanced_search=falseoperation=view_or_manage_repositorypage=1async=falseshow_item_checkboxes=falsef-free-text-search=recalibrationid=797c55906a13241a But I am only a user of usegalaxy public server, so I am afraid I think I can't install tool from tool shed. What I am doing is here: https://usegalaxy.org/u/ksfk/h/illumina-exome I think I need Table Recalibration in NGS: GATK Tools (beta) tool group of left pane of usegalaxy, but I cannot find it. I am wondering whether there is any way to directly call Table Recalibration without web-application wrapper. Regards, Kaz 2014-04-12 7:26 GMT+09:00 Björn Grüning bjoern.gruen...@gmail.com: Hi Kaz, you are searching for GATK tools, right? Please have a look at the GATK suites in the Tool Shed. Cheers, Bjoern Am 12.04.2014 00:16, schrieb KS: Dear all, I can't find either of Table Recalibration or Base Recalibrator on usegalaxy public server. I am defident this is right place to ask, but seqanswers is clouded with many other pipelines. Is there an update of base recalibration tool on usegalaxy? Best, Kaz ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
Re: [galaxy-user] Base Recalibration tool on usegalaxy
Hi Kaz, We do have a problem with the Table Recalibration tool on Main right now. We’ll let you know when we have a fix. Thanks for using Galaxy, Dan On Apr 11, 2014, at 10:24 PM, KS k...@kyudai.jp wrote: Hi Bjoern, Thank you for the reply. I found Table Recalibration, probably from GATK, on Tool Shed as you informed. http://toolshed.g2.bx.psu.edu/repository/view_repository?sort=nameadvanced_search=falseoperation=view_or_manage_repositorypage=1async=falseshow_item_checkboxes=falsef-free-text-search=recalibrationid=797c55906a13241a But I am only a user of usegalaxy public server, so I am afraid I think I can't install tool from tool shed. What I am doing is here: https://usegalaxy.org/u/ksfk/h/illumina-exome I think I need Table Recalibration in NGS: GATK Tools (beta) tool group of left pane of usegalaxy, but I cannot find it. I am wondering whether there is any way to directly call Table Recalibration without web-application wrapper. Regards, Kaz 2014-04-12 7:26 GMT+09:00 Björn Grüning bjoern.gruen...@gmail.com: Hi Kaz, you are searching for GATK tools, right? Please have a look at the GATK suites in the Tool Shed. Cheers, Bjoern Am 12.04.2014 00:16, schrieb KS: Dear all, I can't find either of Table Recalibration or Base Recalibrator on usegalaxy public server. I am defident this is right place to ask, but seqanswers is clouded with many other pipelines. Is there an update of base recalibration tool on usegalaxy? Best, Kaz ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/ ___ 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/ To search Galaxy mailing lists use the unified search at: http://galaxyproject.org/search/mailinglists/
