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I need to merge my all transcripts.gtf from cufflinks output follow this command line :

cuffmerge -o merged_gtf_output -p 15 -s ref.fasta -g anot.gtf assembly.txt

But I have this warning :

[bam_header_read] EOF marker is absent. The input is probably truncated.
[bam_header_read] invalid BAM binary header (this is not a BAM file).
File merge_B3//tmp/mergeSam_fileOaNffV doesn't appear to be a valid BAM file, trying SAM...
[21:40:04] Loading reference annotation.
[21:40:05] Inspecting reads and determining fragment length distribution.
Processed 3921 loci.                        
> Map Properties:
>       Normalized Map Mass: 33242.00
>       Raw Map Mass: 33242.00
>       Fragment Length Distribution: Truncated Gaussian (default)
>                     Default Mean: 200
>                  Default Std Dev: 80

Can someone tell me what this warning means and how to fix it?

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  • $\begingroup$ it looks very weird to me, since you are not reading a bam file, some how samtools is called and it throws this warning, see also biostars.org/p/189139 $\endgroup$
    – StupidWolf
    Mar 29 '20 at 11:26
  • $\begingroup$ Yes it's true that it's weird, but I think it alludes to the files from which the gtf were generated by cufflinks $\endgroup$
    – Diango
    Mar 30 '20 at 12:18
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There is a post about this on seqanswers. The poster says the following about cuffmerge behavior:

I kept the temporary files while running cuffmerge. It builgs a SAM file from transcripts.gtf files. Then it seems first it reads it as BAM since I read samtools take the files as BAM default. Then it finds it is not a BAM so it tries SAM. Then it can read it and resume the task.

So it sounds like the BAM file is generated from GTF as a kludge inside cuffmerge, not the other way around.

Did you change any of the intermediate files or delete anything? Is there maybe a different cuffmerge run that it might be colliding with?

Did you get any output? it sounds like this error is frequently observed when actually the command completes fine, e.g. https://www.biostars.org/p/189139/. See other posts by googling the problem maybe, e.g. https://www.biostars.org/p/344439/

(I know this isn't a proper answer but it was too involved for a comment.)

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  • $\begingroup$ Thank you for your answer, No I did not delete anything and despite the warning the command was executed well. $\endgroup$
    – Diango
    Apr 1 '20 at 11:49

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