Hi Gunter, this is for each a pileup position regardless of the start coordinate.
There is an expensive step in the calculation which is solved by keeping a pre-calculating table for number of reads up to 255. This constraint cannot be easily removed, but 255 reads should be plenty for estimating the genotype. Do the qualities come out vastly different? Please see also the section 3.1.2 and 3.1.3 of the samtools math notes if you are interested in more details http://www.broadinstitute.org/gatk/media/docs/Samtools.pdf petr On Wed, 2016-02-03 at 17:05 +0100, Günter Jäger wrote: > Dear Petr, > > the average coverage of my files varies between 300 and 2000 (very deep > sequencing, paired-end). Now it is clear why I get different results. > However, it is not clear to me why there is this constraint. Or did you > mean max 255 reads mapping to the same start position, but in total more > than 255 covering a candidate site? > > - Is there a way to increase the number of sampled reads for genotype > likelihood estimation, or is this a fixed value? > > Many thanks. > > Günter > > Am 03.02.2016 um 13:09 schrieb Petr Danecek: > > What is the coverage of your bams? samtools samples randomly 255 reads > > when calculating genotype likelihoods. > > > > petr > > > > On Mon, 2016-02-01 at 09:35 -0500, Thomas W. Blackwell wrote: > >> See option -d, --max-depth. Also -F, -L, -m for indel calling. > >> > >> These count all reads overlapping a candidate variant site, rather > >> than by starting position. > >> > >> - tom blackwell - > >> > >> On Mon, 1 Feb 2016, Gnter Jger wrote: > >> > >>> Dear Samtools-Team, > >> I encountered that samtools mpileup followed by bcftools for variant > >> calling produces different results when the bam file is sorted > >> differently. Since the bam file has to be sorted by coordinate, the > >> order of reads sharing the same start position may vary in different > >> versions of the bam file, i.e. if reads in a coordinate sorted bam file > >> are shuffled and then sorted by coordinate again, the order of reads > >> mapping to identical start positions usually differs from the order of > >> the same reads in the initial bam file. > >> > >> This however, does change the quality of the variants that are called by > >> bcftools. Is suspect that the genotype likelihoods change. I would have > >> expected that changing the order of reads mapping to the same start > >> position does not influence the variant calling, when using the exactly > >> same samtools mpileup/bcftools commands. I use the current version 1.3., > >> but also older versions show that problem. > >> > >> Is there any explanation why this is the case? The problem occurs with > >> every bam file I tested so far. It seems like samtools mpileup does not > >> use all of the reads overlapping a variant position for likelihood > >> estimation?! Is there any option in bcftools or samtools mpileup to > >> prevent this from happening, i.e. to produce the same variant quality > >> scores/genotype likelihood values regardless of the order of reads? > >> > >> I would appreciate any comments on that. > >> > >> Best regards, > >> Günter > >> > >> ------------------------------------------------------------------------------ > >> Site24x7 APM Insight: Get Deep Visibility into Application Performance > >> APM + Mobile APM + RUM: Monitor 3 App instances at just $35/Month > >> Monitor end-to-end web transactions and take corrective actions now > >> Troubleshoot faster and improve end-user experience. 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