Re: [galaxy-user] Trouble with RNAseq analysis
Cristian, The ">" is a formatting character; what needs to match is the string after the ">" in the genome file and the entries in the contig column of your GTF. Your GTF is quite different that your genome file; your genome file has 10 contigs labeled by number, but your GTF has many, many contig names labelled by numbers and names. For Cufflinks to work, you can either (a) turn off bias correction or (b) restrict entries in your GTF to those that match your reference genome. Finally, please reply all to emails so that all emails remain on list for archival and community purposes. Thanks, J. On Mar 30, 2011, at 12:02 PM, Cristian Rojas wrote:
Thanks Jeremy. But in genomes fasta files very often any chromosome represents a sequence followed by ">". Then, it is no possible match contig names in GTF with ">" names in Genome fasta. What must I do? Cristian
----- Mensaje original ---- De: Jeremy Goecks <jeremy.goecks@emory.edu> Para: Cristian Rojas <cristianrojasbr@yahoo.com.ar> CC: galaxy-user@lists.bx.psu.edu Enviado: miércoles, 30 de marzo, 2011 12:53:50 Asunto: Re: [galaxy-user] Trouble with RNAseq analysis
Cristian,
The contig names in your GTF file don't match those in your reference (fasta) file. In order for Cufflinks to use a reference GTF, its contigs names must match those in your reference genome.
Best, J.
On Mar 30, 2011, at 11:31 AM, Cristian Rojas wrote:
Thanks Jeremy. I did it. Cristian
----- Mensaje original ---- De: Jeremy Goecks <jeremy.goecks@emory.edu> Para: Cristian Rojas <cristianrojasbr@yahoo.com.ar> CC: galaxy-user@lists.bx.psu.edu Enviado: miércoles, 30 de marzo, 2011 12:02:47 Asunto: Re: [galaxy-user] Trouble with RNAseq analysis
Cristian,
Please share your history with me (History Options --> Share/Publish --> Share
with User --> my email) and I'll take a look.
Thanks, J.
On Mar 30, 2011, at 10:48 AM, Cristian Rojas wrote:
Hi everybody, I am trying to analyze the differential expression between two RNAseq samples. But I found many troubles aligning my reads. I will describe what I did. First I groomed the FastQ files (2). Then I uploaded the Sorghum genome and aligned the reads to it with Tophat. Aftter that, I tried to use Cufflink with the BAM file of Tophat, using as annotation file an uploaded GTF
file and the Sorghum genome, but I received an error message in the three outputs of Cufflink. I tried to align against new brand Maize genome (now at Galaxy), and the same messages. I also converted the BAM file to SAM, but the
same. Any advice? What was wrong? Thanks in advance. Cristian
___________________________________________________________ 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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Hi Jeremy, I turned off the bias correction and didnt use the genome option, just put the refernde (GTF file), but still the problem. Any help? ----- Mensaje original ---- De: Jeremy Goecks <jeremy.goecks@emory.edu> Para: Cristian Rojas <cristianrojasbr@yahoo.com.ar> CC: galaxy-user@lists.bx.psu.edu Enviado: miércoles, 30 de marzo, 2011 14:25:00 Asunto: Re: [galaxy-user] Trouble with RNAseq analysis Cristian, The ">" is a formatting character; what needs to match is the string after the ">" in the genome file and the entries in the contig column of your GTF. Your GTF is quite different that your genome file; your genome file has 10 contigs labeled by number, but your GTF has many, many contig names labelled by numbers and names. For Cufflinks to work, you can either (a) turn off bias correction or (b) restrict entries in your GTF to those that match your reference genome. Finally, please reply all to emails so that all emails remain on list for archival and community purposes. Thanks, J. On Mar 30, 2011, at 12:02 PM, Cristian Rojas wrote:
Thanks Jeremy. But in genomes fasta files very often any chromosome represents a
sequence followed by ">". Then, it is no possible match contig names in GTF with
">" names in Genome fasta. What must I do? Cristian
----- Mensaje original ---- De: Jeremy Goecks <jeremy.goecks@emory.edu> Para: Cristian Rojas <cristianrojasbr@yahoo.com.ar> CC: galaxy-user@lists.bx.psu.edu Enviado: miércoles, 30 de marzo, 2011 12:53:50 Asunto: Re: [galaxy-user] Trouble with RNAseq analysis
Cristian,
The contig names in your GTF file don't match those in your reference (fasta) file. In order for Cufflinks to use a reference GTF, its contigs names must match those in your reference genome.
Best, J.
On Mar 30, 2011, at 11:31 AM, Cristian Rojas wrote:
Thanks Jeremy. I did it. Cristian
----- Mensaje original ---- De: Jeremy Goecks <jeremy.goecks@emory.edu> Para: Cristian Rojas <cristianrojasbr@yahoo.com.ar> CC: galaxy-user@lists.bx.psu.edu Enviado: miércoles, 30 de marzo, 2011 12:02:47 Asunto: Re: [galaxy-user] Trouble with RNAseq analysis
Cristian,
Please share your history with me (History Options --> Share/Publish --> Share
with User --> my email) and I'll take a look.
Thanks, J.
On Mar 30, 2011, at 10:48 AM, Cristian Rojas wrote:
Hi everybody, I am trying to analyze the differential expression between two
RNAseq samples. But I found many troubles aligning my reads. I will describe
what I did. First I groomed the FastQ files (2). Then I uploaded the Sorghum
genome and aligned the reads to it with Tophat. Aftter that, I tried to use Cufflink with the BAM file of Tophat, using as annotation file an uploaded GTF
file and the Sorghum genome, but I received an error message in the three outputs of Cufflink. I tried to align against new brand Maize genome (now at
Galaxy), and the same messages. I also converted the BAM file to SAM, but the
same. Any advice? What was wrong? Thanks in advance. Cristian
___________________________________________________________ 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:
On Mar 30, 2011, at 3:01 PM, Cristian Rojas wrote:
Hi Jeremy, I turned off the bias correction and didnt use the genome option, just put the refernde (GTF file), but still the problem.
Any help?
I was able to run Cufflinks successfully with bias correction turned off; see the history that I've shared with you. Please try one more time and, if it doesn't work, report it using the bug icon next to the broken dataset. Thanks, J.
I tried agaian and the same problem. I tuned off the bias correction but mantained the GFT file. May be this is the problem? I didnt find your history. Thanks ----- Mensaje original ---- De: Jeremy Goecks <jeremy.goecks@emory.edu> Para: Cristian Rojas <cristianrojasbr@yahoo.com.ar> CC: galaxy-user@lists.bx.psu.edu Enviado: miércoles, 30 de marzo, 2011 16:16:14 Asunto: Re: [galaxy-user] Trouble with RNAseq analysis On Mar 30, 2011, at 3:01 PM, Cristian Rojas wrote:
Hi Jeremy, I turned off the bias correction and didnt use the genome option, just put the refernde (GTF file), but still the problem.
Any help?
I was able to run Cufflinks successfully with bias correction turned off; see the history that I've shared with you. Please try one more time and, if it doesn't work, report it using the bug icon next to the broken dataset. Thanks, J.
I tried agaian and the same problem. I tuned off the bias correction but mantained the GFT file. May be this is the problem? I didnt find your history. Thanks
Look for the history I've shared in History Options --> Histories Shared with Me. As requested, if you're still having problems, please report the problematic dataset by clicking on the bug icon. Thanks, J.
participants (2)
-
Cristian Rojas -
Jeremy Goecks