[BioC] Amplicon and exon level read counts and GC content

[Yu Chuan Tai]

Hi Martin,

More questions on your approaches below. If my BAM files are generated by 
Bowtie2 on pair-end fastq files, for scanBamFlag(), should I set 
isPaired=TRUE? Do I need to worry about other input arguments for 
scanBamFlag() or ScanBamParam(), if I want to calculate coverage properly?

Also, summarizeOverlaps() doesn't seem to handle paired-end reads. How to 
get around this, or it won't affect coverage calculation?

Finally, is there any way to calculate base-specific coverage at any 
genomic locus or interval in Rsamtools?
Thanks!

Best,
Yu Chuan

> More specifically, after
>
>  library(Rsamtools)
>  example(scanBam)  # defines 'fl', a path to a bam file
>
> for a _single_ genomic range
>
>  param = ScanBamParam(what="seq",
>                       which=GRanges("seq1", IRanges(100, 500)))
>  dna = scanBam(fl, param=param)[[1]][["seq"]]
>  length(dna)  # 365 reads overlap region
>  alphabetFrequency(dna, collapse=TRUE, baseOnly=TRUE) # 2838 + 3003 GC
>
> though you'd likely want to specify several regions (vector arguments to 
> GRanges) and think about flags (scanBamFlag() and the flag argument to 
> ScanBamParam), read mapping quality, reads overlapping more than one region, 
> etc. (summarizeOverlaps implements several counting strategies, but it is 
> 'easy' to implement arbitrary approaches).
>
>> 
>> Martin
>> 
>>> 
>>> Thanks for any input!
>>> 
>>> Best,
>>> Yu Chuan
>>> 
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>> 
>> 
>
>
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