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Old 01-20-2015, 07:39 AM   #1
Location: paris

Join Date: Jan 2013
Posts: 10
Default generating pileup of paired end fragments

Dear all,
i'm working with paired end rnaseq data. i wanted to generate a pileup with samtools mpileup but noticed that only the mapping reads get counted, and not the unmapped part between two mates, like so:

______READ1--------READ2________ <- mapping
______1 1 1 1 0 0 0 1 1 1 1________ <- pileup

is generating pileups using the whole fragment (read + mate + part in between them) frowned upon for some reason? would it be correct to do it? if yes are there any tools?

alternatively, is there a tool to merge mates and generate the complete fragment in a sam file?

Thank you!

Last edited by Slacanch; 01-20-2015 at 07:42 AM.
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Old 01-20-2015, 07:55 AM   #2
Devon Ryan
Location: Freiburg, Germany

Join Date: Jul 2011
Posts: 3,480

What would a pileup of the uncovered region between mates even look like. They couldn't contribute any sequence, so what would you show for them? The only conceivable reason I could think of to do this is in peak calling for chip-seq or something like that, but then you don't care about the actual sequence so there'd be no point in going to full pileup route.
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Old 01-20-2015, 08:20 AM   #3
Location: paris

Join Date: Jan 2013
Posts: 10

well i don't really care about the sequence, it's for profile comparison.
as long as the positions are right, i'm ok with it.

maybe i shouldn't have called it pileup, but coverage. just a measure of how many reads overlap each position, in order to be able to compare profiles at certain loci (such as transcriptional readthrough for example).

for future reference, i discovered a script that does more or less this in the "misc" folder of samtools. it's called

i'll check it more in detail and report back.
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fragment, paired end, pileup

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