Hello Mervyn,
I just replied to your submission to the bugs list.
For all in general - alternate mapping tools can be used but which are
appropriate depend on the inputs. If the transcriptome is unspliced
(bacterial, etc), then Bowtie is often used. Just be sure to sort the
resulting file before submitting to Cufflinks. If the transcriptome is
spliced, then the mapper should map spliced reads, annotate them so that
this tool suite recognizes them (the "XS attribute"), then the file
sorted before submitting to Cufflinks.
Tophat does all of this by design for spliced reads. If you run the
mapping jobs outside of Galaxy, be aware of how important the reference
genome and any related annotation files (GFF, GTF) planned to be used
downstream are to the success of the results. All inputs must be an
/exact match/. These wiki sections explain:
http://wiki.galaxyproject.org/Support#Reference_genomes
http://wiki.galaxyproject.org/Support#Tools_on_the_Main_server
http://wiki.galaxyproject.org/Support#Custom_reference_genome (best for
smaller genomes, for larger genomes try a cloud Galaxy with scaled
resources for best performance)
Best,
Jen
Galaxy team
On 10/16/13 12:56 AM, Thomas, Mervyn G. (Dr.) wrote:
Hi All,
I am trying to analyse some human RNA-seq dataset which is in BAM
format to generate FPKM values using cufflinks. The ensembl group have
aligned the data using BWA 0.5.9 to generate these BAM datasets. I
realise that TopHat alignments are optimal for Cufflinks, but is it
possible for me to use cufflinks or some other tool to derive FPKM
values from BWA aligned reads?
I tried using cufflinks on one of the BAM files but got an error:
Error running cufflinks.
return code = -11
Command line:
cufflinks -q --no-update-check -I 300000 -F 0.100000 -j 0.150000 -p 8
/galaxy-repl/main/files/006/853/dataset_6853120.dat
The additional output was:
cufflinks v2.1.1
cufflinks -q --no-update-check -I 300000 -F 0.100000 -j 0.150000 -p 8
I have sent it to the galaxy team and awaiting their reply. Any
thoughts about how I can go about analysing this data.
Best wishes
Mervyn
Dr. Mervyn G Thomas BSc, MBChB, PhD
Academic Foundation Doctor
School of Medicine
University of Leicester
UK
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http://galaxyproject.org
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