Problem With Sam-To-Bam Conversion

Question: Problem With Sam-To-Bam Conversion

5.3 years ago by

United Kingdom

Hello, I am trying to map Illumina paired-ends reads from the nematode Caenorhabditis briggsae. Here are the steps of my history: 1)I uploaded and groomed two .fq files corresponding to paired-end reads. In the attributes, I choose the cb4 database. 2)I mapped them using BWA (BWA Version: 0.5.9-r16 BWA run on paired- end data). However, I used the cb3 database here, since the cb4 was not available. 3)I filtered the output SAM file for mapped reads (galaxy tool version 1.0.0). cb3 is still said to be the reference database in the filtered SAM I got as output. 4) Then I tried to convert the SAM into BAM. Here it failed. When I chose the parameters of this tool, what I had for the 1st option "Choose the source for the reference list" is either: -from the history -locally cached I chose locally cached. Then, when running the tool, and I had this message error: "20: SAM-to-BAM on data 16: converted BAM error An error occurred with this dataset: Samtools Version: 0.1.18 (r982:295) No sequences are available for build (cb3), request them by reporting this error." As recommended, I report you now this error ! I found a similar problem in the Galaxy_user mailing list... But I did not get how the problem was solved ! http://user.list.galaxyproject.org/SAM-to-BAM-conversion-problem- td4135498.html I tried to change the attributes of the SAM and filtered-SAM files to cb4, but it did not work. Note also that in the error report, the version is said to be 0.1.18. But when I click on the SAM-to-BAM tool, the header of the page says: SAM-to-BAM (version 1.1.2). I don't know if this matters or not. Could you please help me fixing this issue ? Many thanks ! -- Fabrice Besnard Institute of Biology of the Ecole Normale Supérieure (IBENS) 46 rue d'Ulm, 75230 Paris cedex 05, France 8th floor. Office: Room 802. Lab: Room 817. mail: fbesnard@biologie.ens.fr Tel: +33-1-44-32-39-44

bwa alignment samtools bam • 1.5k views

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modified 5.3 years ago by Politz, Samuel M. • 70 • written 5.3 years ago by Fabrice BESNARD • 130

5.3 years ago by

Politz, Samuel M. • 70

Politz, Samuel M. • 70 wrote:

Hi Fabrice, I work on C. elegans too, and I have found that you need to have the same ce genome database for all steps; otherwise, some tools won't work later in your workflow. With some help from the Galaxy staff, I discovered that there is a version of ce10 in the CloudMap shared information on the Galaxy server. You can find it under "CloudMap ot266 proof of principle dataset". I found it works well to copy this file into your history. When I used GATK tools, it requests the location of your genome datafile: locally cached versus history. If you choose history, it always finds the ce10 version that you put there. Not sure this will help with SAM to BAM conversion, but ce10 is a more up to date version of the database anyway. And the CloudMap version is sorted in a way that is "GATK-friendly", which was necessary for my application. Hope this helps, Sam Politz To: galaxy-user@lists.bx.psu.edu Subject: [galaxy-user] Problem with SAM-to-BAM conversion Hello, I am trying to map Illumina paired-ends reads from the nematode Caenorhabditis briggsae. Here are the steps of my history: 1)I uploaded and groomed two .fq files corresponding to paired-end reads. In the attributes, I choose the cb4 database. 2)I mapped them using BWA (BWA Version: 0.5.9-r16 BWA run on paired- end data). However, I used the cb3 database here, since the cb4 was not available. 3)I filtered the output SAM file for mapped reads (galaxy tool version 1.0.0). cb3 is still said to be the reference database in the filtered SAM I got as output. 4) Then I tried to convert the SAM into BAM. Here it failed. When I chose the parameters of this tool, what I had for the 1st option "Choose the source for the reference list" is either: -from the history -locally cached I chose locally cached. Then, when running the tool, and I had this message error: "20: SAM-to-BAM on data 16: converted BAM error An error occurred with this dataset: Samtools Version: 0.1.18 (r982:295) No sequences are available for build (cb3), request them by reporting this error." As recommended, I report you now this error ! I found a similar problem in the Galaxy_user mailing list... But I did not get how the problem was solved ! http://user.list.galaxyproject.org/SAM-to-BAM-conversion-problem- td4135498.html I tried to change the attributes of the SAM and filtered-SAM files to cb4, but it did not work. Note also that in the error report, the version is said to be 0.1.18. But when I click on the SAM-to-BAM tool, the header of the page says: SAM-to-BAM (version 1.1.2). I don't know if this matters or not. Could you please help me fixing this issue ? Many thanks ! -- Fabrice Besnard Institute of Biology of the Ecole Normale Supérieure (IBENS) 46 rue d'Ulm, 75230 Paris cedex 05, France 8th floor. Office: Room 802. Lab: Room 817. mail: fbesnard@biologie.ens.fr Tel: +33-1-44-32-39-44 ___________________________________________________________ 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/

ADD COMMENT • link written 5.3 years ago by Politz, Samuel M. • 70

Oh sorry, you are using Cb datatase, so embarrassing. Sorry! But the same approach is good, use the same database for all steps, and try to find a way to put it in your history. Best, Sam To: fbesnard@biologie.ens.fr; galaxy-user@lists.bx.psu.edu Subject: Re: [galaxy-user] Problem with SAM-to-BAM conversion Hi Fabrice, I work on C. elegans too, and I have found that you need to have the same ce genome database for all steps; otherwise, some tools won't work later in your workflow. With some help from the Galaxy staff, I discovered that there is a version of ce10 in the CloudMap shared information on the Galaxy server. You can find it under "CloudMap ot266 proof of principle dataset". I found it works well to copy this file into your history. When I used GATK tools, it requests the location of your genome datafile: locally cached versus history. If you choose history, it always finds the ce10 version that you put there. Not sure this will help with SAM to BAM conversion, but ce10 is a more up to date version of the database anyway. And the CloudMap version is sorted in a way that is "GATK-friendly", which was necessary for my application. Hope this helps, Sam Politz To: galaxy-user@lists.bx.psu.edu Subject: [galaxy-user] Problem with SAM-to-BAM conversion Hello, I am trying to map Illumina paired-ends reads from the nematode Caenorhabditis briggsae. Here are the steps of my history: 1)I uploaded and groomed two .fq files corresponding to paired-end reads. In the attributes, I choose the cb4 database. 2)I mapped them using BWA (BWA Version: 0.5.9-r16 BWA run on paired- end data). However, I used the cb3 database here, since the cb4 was not available. 3)I filtered the output SAM file for mapped reads (galaxy tool version 1.0.0). cb3 is still said to be the reference database in the filtered SAM I got as output. 4) Then I tried to convert the SAM into BAM. Here it failed. When I chose the parameters of this tool, what I had for the 1st option "Choose the source for the reference list" is either: -from the history -locally cached I chose locally cached. Then, when running the tool, and I had this message error: "20: SAM-to-BAM on data 16: converted BAM error An error occurred with this dataset: Samtools Version: 0.1.18 (r982:295) No sequences are available for build (cb3), request them by reporting this error." As recommended, I report you now this error ! I found a similar problem in the Galaxy_user mailing list... But I did not get how the problem was solved ! http://user.list.galaxyproject.org/SAM-to-BAM-conversion-problem- td4135498.html I tried to change the attributes of the SAM and filtered-SAM files to cb4, but it did not work. Note also that in the error report, the version is said to be 0.1.18. But when I click on the SAM-to-BAM tool, the header of the page says: SAM-to-BAM (version 1.1.2). I don't know if this matters or not. Could you please help me fixing this issue ? Many thanks ! -- Fabrice Besnard Institute of Biology of the Ecole Normale Supérieure (IBENS) 46 rue d'Ulm, 75230 Paris cedex 05, France 8th floor. Office: Room 802. Lab: Room 817. mail: fbesnard@biologie.ens.fr Tel: +33-1-44-32-39-44 ___________________________________________________________ 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/

ADD REPLY • link written 5.3 years ago by Politz, Samuel M. • 70

Hello, Thank you Sam for your answer. Unfortunately, it still doesn't work. I tried to create new histories with all the datasets having the same reference database in their attributes/options, with no success. I even tried the two reference genomes available as option of the BWA (cb3full & cb3Canonical), but neither worked: the SAM-to-BM conversion always stopped with the same error message. So I would be very grateful to anyone having another idea to fix this issue ! One possibility I see would be to download the reference genome in my history. Except the fact that it takes a lot of memory space, does anyone think it is a bad idea ? If not, from where can I get the reference genome of C. briggsae (last assembly cb4)? In wormbase, there are various files available, but I don't know which one to pick: ftp://ftp.wormbase.org/pub/wormbase/species/c_briggsae/sequence/genomi c/ Thanks for help and advice, Fabrice -- Fabrice Besnard Institute of Biology of the Ecole Normale Supérieure (IBENS) 46 rue d'Ulm, 75230 Paris cedex 05, France 8th floor. Office: Room 802. Lab: Room 817. mail: fbesnard@biologie.ens.fr Tel: +33-1-44-32-39-44

ADD REPLY • link written 5.3 years ago by Fabrice BESNARD • 130

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