On Thu, Nov 7, 2013 at 5:43 AM, bach le <email@example.com> wrote:
> I would like to ask if any of you have experience with generating
> coverage files from PE ssRNA-Seq* data. I got some discussion around
but it seems
> solved yet.
I've no experience with strand-specific protocols, but it sounds like
strand of the first read in the pair indicates the strand. If that is
please confirm as it would inform our design. As suggested in the
just flip the strand of the second ends and use the code earlier in
thread. GAlignmentPairs makes this easy, the accessors first() and
get you the #1 and #2 reads, respectively, as GAlignments.
> Also, does *countOverlaps* function work with GappedAlignmentPairs
> In that case, will the first or last alignment of each pair be used
> calculating the overlap?
GappedAlignmentPairs is now called GAlignmentPairs. Whenever you want
know whether there is a method for a certain signature, use
to see the list or selectMethod() to retrieve the method object for
signature. In this case:
This means we can call countOverlaps with a GAlignmentPairs and any
(of ranges), including a GRangesList of transcripts. The method works
first converting GAlignmentPairs into a GRangesList, where each
corresponds to a pair and contains the exonic ranges from both ends.
problem for you is that the strand will not be correct for the second
Thus, you will need to call grglist() to get the GRangesList manually
modify the strand before passing it to countOverlaps. The strand
modification would look like:
strand(grl) <- relist(rep(strand(first(reads)), elementLengths(grl)),
There is some experimental functionality in GenomicRanges for
isoform-specific overlaps: findSpliceOverlaps. These functions look
BAM XS tag and if found use it to determine the strand in a way
above. If you were to set the XS mcol on your GAlignmentPairs to
strand(first(reads)), then these functions should work the way you
Thanks much in advance.
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