[Samtools-devel] hard clip before ins/del

[Stuart B. <stu...@pt...>]

Hello,

If I hard-clipped bases from the beginning of reads after alignment, it
leaves me with the possibility of a correctly-mapped deletion or insertion
beginning a read. The SAM spec seems ambiguous to me on whether this is
disallowed, but SAMtools 0.1.18 does not count leading deletions or
insertions in the pileup, it seems. For example, if I hard clip the first
three bases from the following aligned read:

reference: ACTGACGTCA
read: ACTCGTCA
CIGAR: 3M2D5M

After hard-clipping, this is now:

read: CGTCA
CIGAR 3H2D5M

 

I have asked around about how to deal with this, and I frequently get the
response that the aligned read should be considered to begin with the sixth
base of the reference, however when working with amplicons, this deletion
may be critical.  If there is a deletion or insertion after a primer
sequence which gets hard-clipped, all of the reads will have this variant in
the same relative position, and therefore will be missed.  This is important
to me because I usually design primers close to my target region.  I have
made a workaround to check for this condition in my aligner, but I wanted to
make this issue known since I use SAMtools.  Thanks for any advice.

Regards, 

Stuart