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Old 01-29-2014, 12:47 AM   #2
Devon Ryan
Location: Freiburg, Germany

Join Date: Jul 2011
Posts: 3,480

Regarding the negative alignment scores, the highest possible alignment score in end-to-end alignment is 0. For each mismatch or gap open/extension, that score is decremented by some amount (for mismatches, this amount depends on the phred score of the base).

Regarding the MAPQ value, this is due to you using "-k 1". What you're telling bowtie2 to do is to look for at most one valid hit and to stop looking once it's found one. That hit will often not be the best hit that bowtie2 could find and since it's being told to stop looking right away it can't possibly calculate a MAPQ, since it would need to search more thoroughly to have any clue how good the returned alignment is. So, 255 would be the appropriate value.

Just don't use "-k 1" and you'll get the single best alignment for each read and an accompanying MAPQ score.
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