Hello, I was doing a RNA analyse and I wished to compare the transcription and expression of two samples using a reference annotation, however this is the error message I got:
=====Quote===== Error running cuffmerge. [Thu Jul 4 07:32:59 2013] Beginning transcriptome assembly merge -------------------------------------------
[Thu Jul 4 07:32:59 2013] Preparing output location cm_output/ [Thu Jul 4 07:34:07 2013] Converting GTF files to SAM [07:34:07] Loading reference annotation. [07:34:07] Loading reference annotation. [Thu Jul 4 07:34:08 2013] Quantitating transcripts You are using Cufflinks v2.1.1, which is the most recent release. Command line: cufflinks -o cm_output/ -F 0.05 -g /galaxy/main_pool/pool7/files/006/446/dataset_6446730.dat -q --overhang-tolerance 200 --library-type=transfrags -A 0.0 --min-frags-per-transfrag 0 --no-5-extend -p 4 cm_output/tmp/mergeSam_fileIO17rb [bam_header_read] EOF marker is absent. [bam_header_read] invalid BAM binary header (this is not a BAM file). File cm_output/tmp/mergeSam_fileIO17rb doesn't appear to be a valid BAM file, trying SAM... [07:34:08] Loading reference annotation. [07:35:53] Inspecting reads and determining fragment length distribution. Processed 33854 loci.
Map Properties: Normalized Map Mass: 8719.00 Raw Map Mass: 8719.00 Fragment Length Distribution: Truncated Gaussian (default) Default Mean: 200 Default Std Dev: 80
[07:35:53] Assembling transcripts and estimating abundances. Processed 33854 loci. [Thu Jul 4 07:39:29 2013] Comparing against reference file /galaxy/main_pool/pool7/files/006/446/dataset_6446730.dat You are using Cufflinks v2.1.1, which is the most recent release. Error: duplicate GFF ID 'ENST00000361547.2' encountered! [FAILED] Error: could not execute cuffcompare
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The job goes well without the annotation reference. The annotation file I used can be downloaded here: ftp://ftp.sanger.ac.uk/pub/gencode/release_17/gencode.v17.annotation.gtf.gz
Can anyone help me please? Thanks, Delong