Camille -
The samtools command line looks fine except for the final 's1_sortcoords'.
Not sure what's intended there. If this is meant to be the output file
name, then either a redirection operator '>' is needed just before it, or
it should be the value of '-o' and be moved earlier before the input '-'.
I'm not at all familiar with 'subread', but my first step in debugging
would be to run 'samtools view -H' and see whether the .bam header lines
are what you expect them to be. Are the individual reads longer than,
say, 10 kb after assembly ? Does 'subread' expect an additional .bai
file, created with 'samtools index' ? Might try adding that. Just
guessing.
- tom blackwell -
On Sun, 26 Oct 2014, Camille A. Daniels wrote:
> Hello,
>
> I have a bam file formatting issue.
>
> samtools-0.1.18 (Linux) shows the bam to contain 875 reads mapped.
>
>
> [cid:c32e9022-0c2c-4630-85ab-c9d4ce1f4bf9]
>
> I've run featureCounts, a software module of Subread
> (http://subread.sourceforge.net/) to count the number of read mapped per CDS
> to a reference genome.
>
> The program only processes 116 reads for this bam (rather than the expected
> 875), and suggests truncation or missing data.
>
>
> [cid:5abca696-b95f-4911-a199-37a47ebf33db]
>
>
> Can someone help debug the formatting issue?
>
> Workflow to create the bam file is as follows:
> -De novo assembly of reads in CLC Workbench
> -Bowtie2 to map reads against reference genome
> -Pipe to samtools to generate bam and sort file by coordinate
> -Run in featureCounts
>
> ./bowtie2 -x bact_contigs -f -a -t --no-unal -p 36 s1_assembly.fasta |
> ~/samtools-0.1.18/samtools view -Shu - | ~/samtools-0.1.18/samtools sort -m
> 5000000000 - s1_sortcoord
>
> I've re-generated the bam files with and without piping to samtools, and
> versions of both software, but still get the same error.
>
>
> Also, below is a file size comparison for the file and its index.
>
>
> [cid:32f4bea4-956a-4a56-9b26-26c8ef290ecc]
>
>
>
>
>
> ---
> Thanks,
> Camille Daniels, Ph.D.
> Post Doctoral Fellow
> Red Sea Research Center
> Al-Haytham West (Building 2), Room 2227-WS02
> Thuwal, Saudi Arabia 23599-6900
>
> camille.dani...@kaust.edu.sa
> Office: +966 02 808 2446
> ---
>
> ________________________________
>
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