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What could make Bowtie2 miss legitimate hits?

Hi,

We've noticed some strange behavior by Bowtie2: In specific cases - it simply misses a proper hit for a read. We noticed this after investigating unmapped reads - we were surprised to see that they have a perfect end-to-end match to the genome we were aligning them against. This raised the question - why did Bowtie fail to catch this hit.

Repeating the process showed that these were the same reads that bowtie was failing with (so this is not a random error rate). These reads are trimmed, contain no adaptor, have high quality, and proper length, and - as mentioned above - have a perfect match in the genome.

Any idea why Bowtie2 is missing their alignemnt?

Thanks, Iddo

alignment
• 1,052 views • link
written by iddo.nadav • iddo.nadav

One of a set of PCR duplicates? Can you filter them out with picard tools? What does the bam flag say? Did you try a different aligner, e.g. BWA mem?

• 0 views • link
written by Carambakaracho Carambakaracho

Yes, we did. Surprisingly BWA was able to map these reads. How would you interpret this?

• 0 views • link
written by iddo.nadav • iddo.nadav

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