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  • cuffmerge error

    Hi, All

    I was trying to run the cuffmerge for my two samples. However, there is warning message point to the mitochondrial genome, however, i could not figure out the exact problem, could you please give some hints? I directly download the Saccharomyces cerevisiae/Ensembl/EF4, ref genome.

    Thank you very much!

    bq@bq-VirtualBox:~/Desktop/rnaseq/trimmed$ cuffmerge -g genes.gtf -s genome.fa -p 4 assemblies.txt

    [Wed Jun 5 22:46:37 2013] Beginning transcriptome assembly merge
    -------------------------------------------

    [Wed Jun 5 22:46:37 2013] Preparing output location ./merged_asm/
    [Wed Jun 5 22:46:37 2013] Converting GTF files to SAM
    [22:46:37] Loading reference annotation.
    [22:46:37] Loading reference annotation.
    [Wed Jun 5 22:46:37 2013] Quantitating transcripts
    You are using Cufflinks v2.1.1, which is the most recent release.
    Command line:
    cufflinks -o ./merged_asm/ -F 0.05 -g genes.gtf -q --overhang-tolerance 200 --library-type=transfrags -A 0.0 --min-frags-per-transfrag 0 --no-5-extend -p 4 ./merged_asm/tmp/mergeSam_fileDalcVi
    [bam_header_read] EOF marker is absent.
    [bam_header_read] invalid BAM binary header (this is not a BAM file).
    File ./merged_asm/tmp/mergeSam_fileDalcVi doesn't appear to be a valid BAM file, trying SAM...
    [22:46:37] Loading reference annotation.
    [22:46:38] Inspecting reads and determining fragment length distribution.
    Processed 2290 loci.
    > Map Properties:
    > Normalized Map Mass: 6966.00
    > Raw Map Mass: 6966.00
    > Fragment Length Distribution: Truncated Gaussian (default)
    > Default Mean: 200
    > Default Std Dev: 80
    [22:46:38] Assembling transcripts and estimating abundances.
    Processed 2290 loci.
    [Wed Jun 5 22:46:43 2013] Comparing against reference file genes.gtf
    You are using Cufflinks v2.1.1, which is the most recent release.
    Warning: couldn't find fasta record for 'Mito'!
    [Wed Jun 5 22:46:44 2013] Comparing against reference file genes.gtf
    You are using Cufflinks v2.1.1, which is the most recent release.
    Warning: couldn't find fasta record for 'Mito'!

  • #2
    The fasta sequence for "Mito" is missing from the genome.fa file.

    Comment


    • #3
      Hi, GenoMax

      I should have mentioned that i did some initial troubleshooting, you are right that "Mito" is missing in the genome.fa file. It was named as "MT" instead. So I renamed this file, and reran it, the same error occurred again. Maybe something else has to be done besides renaming, or it is a totally different thing?

      Best,

      Comment


      • #4
        So the chromosome names in your genes.gtf are not matching what you had in the original reference file (your alignments inherited those names).

        Compare these two commands:
        Code:
        awk -F "\t" '{print $1}' genes.gtf | uniq
        and

        Code:
        cat genome.fa | grep ">"

        If that is true then you could rename the "Mito" from the genes.gtf to "MT" and then try a re-run.
        Last edited by GenoMax; 06-06-2013, 08:53 AM.

        Comment


        • #5
          Thank you so much! The error does go away when I rename the "mito" to "MT". It just never came to me that the two files' name are inconsistent. Really appreciate your help!

          Baoqing

          Comment

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