Re: [galaxy-user] Cuffdiff question
Hi Noa, Yes I did use Cufflinks so this sounds just like my problem. So how have you dealt with the problem? Best wishes, Clare On 14 Nov 2013, at 18:17, Noa Sher wrote: > Hi Clare > We just ran into a similar issue about a week ago and were debugging with the > authors of cuffdiff > Apparently there are issues with the -b parameter - were you using this in > cufflinks? > If yes - this may be the cause - we switched the order of the replicates and > the values changed; as did PCA's of the samples, etc > They are working on this issue for an upcoming version of cufflinks. > I am interested in knowing whether this was indeed your problem or were you > using a pipeline that does not include cufflinks? > Good luck, > Noa > > > On 14/11/2013 13:13, clare Hardman wrote: >> Hello, >> >> Could you please advise me on this probably naive question. When I compare >> sample A and sample B by Ciffdiff and then separately compare Sample A to >> Sample C by Cuffdiff too, should the FMPK value be the same for A in both >> tests? At the moment mine does not seem to be! >> >> Best wishes >> >> Clare >> >> >> ___ >> 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] Cuffdiff question
Dont use the - b parameter Sent from my iPhone; please excuse any brevity or typos! On Nov 15, 2013, at 2:51 PM, clare Hardman wrote: > Hi Noa, > > Yes I did use Cufflinks so this sounds just like my problem. So how have you > dealt with the problem? > > Best wishes, > > Clare > > > > On 14 Nov 2013, at 18:17, Noa Sher wrote: > >> Hi Clare >> We just ran into a similar issue about a week ago and were debugging with >> the authors of cuffdiff >> Apparently there are issues with the -b parameter - were you using this in >> cufflinks? >> If yes - this may be the cause - we switched the order of the replicates and >> the values changed; as did PCA's of the samples, etc >> They are working on this issue for an upcoming version of cufflinks. >> I am interested in knowing whether this was indeed your problem or were you >> using a pipeline that does not include cufflinks? >> Good luck, >> Noa >> >> >> On 14/11/2013 13:13, clare Hardman wrote: >>> Hello, >>> >>> Could you please advise me on this probably naive question. When I compare >>> sample A and sample B by Ciffdiff and then separately compare Sample A to >>> Sample C by Cuffdiff too, should the FMPK value be the same for A in both >>> tests? At the moment mine does not seem to be! >>> >>> Best wishes >>> >>> Clare >>> >>> >>> ___ >>> 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] Cuffdiff question
Hi Clare We just ran into a similar issue about a week ago and were debugging with the authors of cuffdiff Apparently there are issues with the -b parameter - were you using this in cufflinks? If yes - this may be the cause - we switched the order of the replicates and the values changed; as did PCA's of the samples, etc They are working on this issue for an upcoming version of cufflinks. I am interested in knowing whether this was indeed your problem or were you using a pipeline that does not include cufflinks? Good luck, Noa On 14/11/2013 13:13, clare Hardman wrote: Hello, Could you please advise me on this probably naive question. When I compare sample A and sample B by Ciffdiff and then separately compare Sample A to Sample C by Cuffdiff too, should the FMPK value be the same for A in both tests? At the moment mine does not seem to be! Best wishes Clare ___ 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] Cuffdiff question about using an unspecified (?) database/build
Jen, Thank you very much for the reply. I'm glad to know it is a known bug and not something on my side of things. So, would my analysis be affected if I did change the bam file "Database/Build" to the older tree shrew version found in the drop down list? What significance does this "Database/Build" box have in downstream analysis if you have your own fasta reference genome file and gtf annotation file that is being referenced instead of a locally cached one? I'm just trying to obtain a better understanding of the "Database/Build" box for analyses where I provide the fasta and gtf file. Thanks, David -Original Message- From: Jennifer Jackson [mailto:j...@bx.psu.edu] Sent: Friday, August 19, 2011 9:20 AM To: David K Crossman Cc: galaxy-user (galaxy-user@lists.bx.psu.edu) Subject: Re: [galaxy-user] Cuffdiff question about using an unspecified (?) database/build Hello David, This is a known bug. The correction is planned to be moved out onto the public Galaxy instance at the next update (within a week). Sorry for the current inconvenience, Best, Jen Galaxy team On 8/19/11 7:00 AM, David K Crossman wrote: > Hello! > > I have an RNA-Seq project which consists of 5 samples from the species > tree shrew. When uploading these fastq files into Galaxy, I chose > "unspecified (?)" for the database/build since the latest tree shrew > version is not in the drop down list. When using TopHat, > Cufflinks/Compare I have selected a reference genome from my history > instead of using a built-in index, as well as a gtf annotation file > for Cufflinks/Compare and everything has been working fine. Now, I am > at the Cuffdiff step and I am running into an error when setting it up > to perform replicate analysis. When I select my TopHat accepted hits > bam file I see a red X and the error: "Unspecified genome build, click > the pencil icon in the history item to set the genome build." Here's a > screenshot of what I'm seeing: > > Since the latest reference genome for tree shrew wasn't listed, that's > why I chose "unspecified (?)." Should I go back and edit these > accepted hits bam files to say the Database/Build from the drop down > list is "Tree shrew Dec. 2006 (Broad/tupBel1) (tupBel1)?" I know that > this is simple to change, but will this affect my results in any way? > Any help/info would be greatly appreciated. > > Thanks, > > David > > > > ___ > 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://usegalaxy.org http://galaxyproject.org/Support ___ 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/
Re: [galaxy-user] Cuffdiff question about using an unspecified (?) database/build
Hello David, This is a known bug. The correction is planned to be moved out onto the public Galaxy instance at the next update (within a week). Sorry for the current inconvenience, Best, Jen Galaxy team On 8/19/11 7:00 AM, David K Crossman wrote: Hello! I have an RNA-Seq project which consists of 5 samples from the species tree shrew. When uploading these fastq files into Galaxy, I chose “unspecified (?)” for the database/build since the latest tree shrew version is not in the drop down list. When using TopHat, Cufflinks/Compare I have selected a reference genome from my history instead of using a built-in index, as well as a gtf annotation file for Cufflinks/Compare and everything has been working fine. Now, I am at the Cuffdiff step and I am running into an error when setting it up to perform replicate analysis. When I select my TopHat accepted hits bam file I see a red X and the error: “Unspecified genome build, click the pencil icon in the history item to set the genome build.” Here’s a screenshot of what I’m seeing: Since the latest reference genome for tree shrew wasn’t listed, that’s why I chose “unspecified (?).” Should I go back and edit these accepted hits bam files to say the Database/Build from the drop down list is “Tree shrew Dec. 2006 (Broad/tupBel1) (tupBel1)?” I know that this is simple to change, but will this affect my results in any way? Any help/info would be greatly appreciated. Thanks, David ___ 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://usegalaxy.org http://galaxyproject.org/Support ___ 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/
Re: [galaxy-user] Cuffdiff Question
Thanks for the reply. I tried to use the script provided on a previous galaxy thread for adding the chr on to the gtf file on the mac terminal but I keep getting this error - awk: can't open file ensembl.gtf source line number 1 I am very new to using the terminal so please let me know if there is something basic that I am not doing right, Thanks! Kurinji On Tue, Jun 28, 2011 at 6:13 AM, Jeremy Goecks wrote: > Hello Kurinji, > > I was at your USC Galaxy seminar last week, which I found very helpful - > thank you! > > > Glad to hear that you found the workshop helpful. As a reminder, please > email questions about using Galaxy and its tools to the galaxy-user mailing > list (which I've cc'd). You may get quicker and different responses from > community members, and everyone will benefit from the discussion. > > I used my recently generated RNAseq data in Galaxy (which was pre-aligned > using tophat and already had cufflinks run on it) - I ran cuffcompare with > all the gtf files and then cuffdiff for the three pairs (there is 1 control > and 3 different drug treatments - no replicates). I got several output > files, as expected, but decided just to look at the gene differential > expression as a start. Some questions I have are - > > 1. (very basic question!) which is sample 1 (and corresponding value 1) and > sample 2 (and corresponding value 2)in my output file. This is what my > output file is called - > > 90: Cuffdiff on data 37, data 38, and data 60: gene differential expression > testing 33,969 lines > > Is 37 sample one or sample two? Given the data - I would expect sample 37 > to correspond to "value 2" - but I could be wrong. Please let me know! > > > The best way to figure out which dataset corresponds with Cuffdiff's labels > is to click the rerun button in the dataset: sample names correspond > directly to the reads datasets (i.e. BAM files) provided as input to > Cuffdiff. > > 2. How do I find the UCSC gene names corresponding with start/end sites - I > did input the hg18 UCSC gtf file as a reference > > > You'll need to use a reference annotation (GTF file) that has the gene_name > attribute as input for Cufflinks/compare/difff. Typically Ensembl > annotations have this attribute; however, you'll need to prepend 'chr' to > each line--really, to each chromosome name--in order to bring Ensembl > notation in line with UCSC/Galaxy notation. > > Actually, I noticed that value 1 in this particular output file is all 0 - > no idea why. It is not this way in the other files, making me wonder if > there is an error somewhere. I am sure the bam file is okay as I viewed it > on IGV and saw the patterns I would expect for some candidate genes I looked > at. > > > It's difficult for me to comment without seeing your analysis. Some output > files depend on particular attributes being set correctly in the annotation > file. You may want to search through our mailing list archives and see if > your question has already been answered: > http://gmod.827538.n3.nabble.com/Galaxy-Users-f815892.html > > Good luck, > J. > ___ 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/
Re: [galaxy-user] Cuffdiff Question
> Thanks for the reply. I tried to use the script provided on a previous galaxy > thread for adding the chr on to the gtf file on the mac terminal but I keep > getting this error - > > awk: can't open file ensembl.gtf > source line number 1 > > I am very new to using the terminal so please let me know if there is > something basic that I am not doing right, Try this Galaxy workflow: http://main.g2.bx.psu.edu/u/jeremy/w/make-ensembl-gtf-compatible-with-cufflinks It simply prepends 'chr' to the chromosome name, which is needed if you're using an Ensemble reference annotation and want to use it with Cufflinks/compare/diff in Galaxy. Best, J. ___ 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/
Re: [galaxy-user] Cuffdiff Question
Hello Kurinji, > I was at your USC Galaxy seminar last week, which I found very helpful - > thank you! Glad to hear that you found the workshop helpful. As a reminder, please email questions about using Galaxy and its tools to the galaxy-user mailing list (which I've cc'd). You may get quicker and different responses from community members, and everyone will benefit from the discussion. > I used my recently generated RNAseq data in Galaxy (which was pre-aligned > using tophat and already had cufflinks run on it) - I ran cuffcompare with > all the gtf files and then cuffdiff for the three pairs (there is 1 control > and 3 different drug treatments - no replicates). I got several output files, > as expected, but decided just to look at the gene differential expression as > a start. Some questions I have are - > > 1. (very basic question!) which is sample 1 (and corresponding value 1) and > sample 2 (and corresponding value 2)in my output file. This is what my output > file is called - > > 90: Cuffdiff on data 37, data 38, and data 60: gene differential expression > testing 33,969 lines > > Is 37 sample one or sample two? Given the data - I would expect sample 37 to > correspond to "value 2" - but I could be wrong. Please let me know! The best way to figure out which dataset corresponds with Cuffdiff's labels is to click the rerun button in the dataset: sample names correspond directly to the reads datasets (i.e. BAM files) provided as input to Cuffdiff. > 2. How do I find the UCSC gene names corresponding with start/end sites - I > did input the hg18 UCSC gtf file as a reference You'll need to use a reference annotation (GTF file) that has the gene_name attribute as input for Cufflinks/compare/difff. Typically Ensembl annotations have this attribute; however, you'll need to prepend 'chr' to each line--really, to each chromosome name--in order to bring Ensembl notation in line with UCSC/Galaxy notation. > Actually, I noticed that value 1 in this particular output file is all 0 - no > idea why. It is not this way in the other files, making me wonder if there is > an error somewhere. I am sure the bam file is okay as I viewed it on IGV and > saw the patterns I would expect for some candidate genes I looked at. It's difficult for me to comment without seeing your analysis. Some output files depend on particular attributes being set correctly in the annotation file. You may want to search through our mailing list archives and see if your question has already been answered: http://gmod.827538.n3.nabble.com/Galaxy-Users-f815892.html Good luck, J.___ 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/
