[BioC] Filtering BAM files by start position for VariantTools

[Taylor, Sean D]

Thanks Valerie, I'll give that a try. And I see that you just clarified my question about the input file for tallyVariants.

I had thought about doing this before but hadn't been able to figure out the proper filter syntax. Thanks!  The only downside to this approach is that I ultimately want to do this for all unique start sites that align to my reference. That could entail generating thousands of tempfiles that all have to be read back in. It could be done with lapply, but it sounds rather memory and time intensive. Another approach that I tried was reading the bam file as a GappedAlignments object and then splitting it by start position into a GAlignmentsList. That was really easy, but so far as I can tell tallyVariants will not accept GappedAlignments objects as an input. I wonder if there is another way to vectorize this approach?

Thanks,
Sean


-----Original Message-----
From: Valerie Obenchain [mailto:vobencha at fhcrc.org] 
Sent: Thursday, July 11, 2013 10:02 AM
To: Taylor, Sean D
Cc: bioconductor at r-project.org; Michael Lawrence
Subject: Re: [BioC] Filtering BAM files by start position for VariantTools

Hi Sean,

As you've discovered, the 'which' in the 'param' (for reading bam files) specifies positions to overlap, not start or end. One approach to isolating reads with a specific starting position would be to filter the bam by 'pos'.

     library(VariantTools)
     fl <- LungCancerLines::LungCancerBamFiles()$H1993

     mystart <- 1110426
     filt <- list(setStart=function(x) x$pos %in% mystart)
     dest <- tempfile()
     filterBam(fl, dest, filter=FilterRules(filt))
     scn <- scanBam(dest)

Confirm all reads start with 'mystart':

 > table(scn[[1]]$pos)

1110426
    2388

If you want a tally of all nucleotides for all sequences starting with 'mystart' then no need to supply a 'which':
     param <- VariantTallyParam(gmapR::TP53Genome(),
                                readlen=100L,
                                high_base_quality=23L)
     tally <- tallyVariants(fl, param)


Valerie


On 07/09/2013 02:06 PM, Taylor, Sean D wrote:
> I am trying to read a specific set of records from a bam file for use in the VariantTools package. I'm trying to construct a which argument (a GRanges object) that will pull in a set of records from all reads that only start at a specified position. (i.e. all reads that start at position 100). So far I have only been able to specify reads that overlap position 100, but have not been able to find a way to define the start site.
>
> #Example code:
>> bams <- LungCancerLines::LungCancerBamFiles()
>> bam <- bams$H1993
>> which<-GRanges(seqnames=c('TP53'), IRanges(1110426, width=1), '+')
>> tally.param <- VariantTallyParam(gmapR::TP53Genome(),
> + readlen = 100L,
> + high_base_quality = 23L,
> + which = which)
>> raw.variants <- tallyVariants(bam, tally.param)
>
> This code shows all the variants at position 1110426, but not all the variants from the reads that start at position 1110426.
>
> Ultimately, I am trying to do this for all start positions in my data set, so I would want something that looks like this pseudocode:
>> raw.variants<-lapply (start(bam), function (x){
>    which<-GRanges(seqnames=c('chrM'), '+', start=x)
>    tally.param<-VariantTallyParam(gmap, readlen=100L, which=which)
>    tallyVariants(bamfile, tally.param)
> })
>
> Thanks,
> Sean
>
>> sessionInfo()
> R version 3.0.1 (2013-05-16)
> Platform: x86_64-unknown-linux-gnu (64-bit)
>
> locale:
> [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C               LC_TIME=en_US.UTF-8
>   [4] LC_COLLATE=en_US.UTF-8     LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8
>   [7] LC_PAPER=C                 LC_NAME=C                  LC_ADDRESS=C
> [10] LC_TELEPHONE=C             LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
>
> attached base packages:
> [1] grid      parallel  stats     graphics  grDevices utils     datasets  methods   base
>
> other attached packages:
> [1] RSQLite_0.11.4          DBI_0.2-7               BiocInstaller_1.10.2
>   [4] LungCancerLines_0.0.8   GenomicFeatures_1.12.1  AnnotationDbi_1.22.5
>   [7] Biobase_2.20.0          gmapR_1.2.0             latticeExtra_0.6-24
> [10] lattice_0.20-15         RColorBrewer_1.0-5      genoPlotR_0.8
> [13] ade4_1.5-2              VariantTools_1.2.2      VariantAnnotation_1.6.6
> [16] Rsamtools_1.12.3        Biostrings_2.28.0       GenomicRanges_1.12.4
> [19] IRanges_1.18.1          BiocGenerics_0.6.0
>
> loaded via a namespace (and not attached):
> [1] biomaRt_2.16.0                          bitops_1.0-5
>   [3] BSgenome_1.28.0                         BSgenome.Hsapiens.UCSC.hg19_1.3.19
>   [5] graph_1.38.2                            Matrix_1.0-12
>   [7] org.Hs.eg.db_2.9.0                      RBGL_1.36.2
>   [9] RCurl_1.95-4.1                          rtracklayer_1.20.2
> [11] stats4_3.0.1                            tools_3.0.1
> [13] TxDb.Hsapiens.UCSC.hg19.knownGene_2.9.2 XML_3.96-1.1
> [15] zlibbioc_1.6.0
>
>
> 	[[alternative HTML version deleted]]
>
> _______________________________________________
> Bioconductor mailing list
> Bioconductor at r-project.org
> https://stat.ethz.ch/mailman/listinfo/bioconductor
> Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor
>