Hi Jesper,
On 02/19/2014 08:44 AM, Michael Lawrence wrote:
On Wed, Feb 19, 2014 at 8:39 AM, Jesper Gådin <jesper.ga...@gmail.com>wrote:
Hi Michael,
Herves suggestion will probably work for my use case, but if there are any
smoother ways it would be preferable.
The use case is as follows:
1) calculate strand specific coverage over a region from
GAlignments object (or file)
At the moment I read a file using readGAlignmentsFromBam() with tag="XS",
then filter it on "flag" and "mapq". Then I subset the
resulting GAlignments in a minus and a plus -strand object.
Then I calculate coverage by my own coverage function which uses the cigar
information in the GAlignments object. This function is the one using
cigarToRleList() at the moment. As I understand the coverage() function
from the GenomicAlignments/IRanges package does not take into account
cigars, or does it?
It does take the coverage into account; specifically to exclude the introns
from coverage. I think there's also an option to exclude deletions.
Unfortunately the man page is not easy to access (you need to do
?`coverage,GAlignments-method`), but it says:
The methods for GAlignments and GAlignmentPairs objects do:
coverage(grglist(x), ...)
And if you do grglist() on a GAlignments or GAlignmentPairs objects, the
ranges you get in the returned GRangesList object are calculated based
on the CIGAR.
Trust but verify. Here is how you can actually verify that coverage()
does take the CIGAR into account:
library(RNAseqData.HNRNPC.bam.chr14)
gal <- readGAlignments(RNAseqData.HNRNPC.bam.chr14_BAMFILES[1])
cig_op_table <- cigarOpTable(cigar(gal))
First we pick up an alignment with an N in its CIGAR and do coverage()
on it:
> gal_with_N <- gal[which(cig_op_table[ , "N"] != 0)[1]]
> gal_with_N
GAlignments with 1 alignment and 0 metadata columns:
seqnames strand cigar qwidth start end width
<Rle> <Rle> <character> <integer> <integer> <integer> <integer>
[1] chr14 + 55M2117N17M 72 19650072 19652260 2189
ngap
<integer>
[1] 1
---
seqlengths:
chr1 chr10 ... chrY
249250621 135534747 ... 59373566
> coverage(gal_with_N)$chr14
integer-Rle of length 107349540 with 5 runs
Lengths: 19650071 55 2117 17 87697280
Values : 0 1 0 1 0
Same thing with an alignment with an I in its CIGAR:
> gal_with_I <- gal[which(cig_op_table[ , "I"] != 0)[1]]
> gal_with_I
GAlignments with 1 alignment and 0 metadata columns:
seqnames strand cigar qwidth start end width
<Rle> <Rle> <character> <integer> <integer> <integer> <integer>
[1] chr14 - 64M1I7M 72 19411677 19411747 71
ngap
<integer>
[1] 0
---
seqlengths:
chr1 chr10 ... chrY
249250621 135534747 ... 59373566
> coverage(gal_with_I)$chr14
integer-Rle of length 107349540 with 3 runs
Lengths: 19411676 71 87937793
Values : 0 1 0
Same thing with an alignment with a D in its CIGAR:
> gal_with_D <- gal[which(cig_op_table[ , "D"] != 0)[1]]
> gal_with_D
GAlignments with 1 alignment and 0 metadata columns:
seqnames strand cigar qwidth start end width
<Rle> <Rle> <character> <integer> <integer> <integer> <integer>
[1] chr14 + 38M1D34M 72 19659063 19659135 73
ngap
<integer>
[1] 0
---
seqlengths:
chr1 chr10 ... chrY
249250621 135534747 ... 59373566
> coverage(gal_with_D)$chr14
integer-Rle of length 107349540 with 3 runs
Lengths: 19659062 73 87690405
Values : 0 1 0
Seeing is believing,
Cheers,
H.
I started to look at the applyPileups() in Rsamtools which I can get to
calculate coverage using cigars, but not using the strand or flag
information for filtering. That solution would start from a bam-file
instead of a GAlignments object, and sure I can do the filtering outside R.
But it would be very convenient to do it all from within R.
If there are nice solutions starting from both a GAlignments and a
bam-file it would be great! =)
/Jesper
On Tue, Feb 18, 2014 at 10:52 PM, Michael Lawrence <
lawrence.mich...@gene.com> wrote:
Hi Jesper,
Would you be willing to volunteer your use case? As Herve hinted,
cigarToRleList and friends are low-level helpers. There may be an easier
way to achieve what you want, or an opportunity to improve things.
Michael
On Mon, Feb 17, 2014 at 1:10 AM, Jesper Gådin <jesper.ga...@gmail.com>wrote:
Hi,
Have come across a cigar-vector that is problematic to process.
#load package
library(GenomicAlignments)
#load data (see attached file)
load("2014-02-17-cigarExample.rdata")
#run function cigarToRleList
cigarRle <- cigarToRleList(cigarExample)
Error in .Call2("Rle_constructor", values, lengths, check, 0L, PACKAGE =
"IRanges") :
integer overflow while summing elements in 'lengths'
sessionInfo()
R Under development (unstable) (2013-11-13 r64209)
Platform: x86_64-unknown-linux-gnu (64-bit)
locale:
[1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C
[3] LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8
[5] LC_MONETARY=en_US.UTF-8 LC_MESSAGES=en_US.UTF-8
[7] LC_PAPER=en_US.UTF-8 LC_NAME=C
[9] LC_ADDRESS=C LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
attached base packages:
[1] parallel stats graphics grDevices utils datasets methods
[8] base
other attached packages:
[1] GenomicAlignments_0.99.18 Rsamtools_1.15.26
[3] Biostrings_2.31.12 XVector_0.3.6
[5] GenomicRanges_1.15.26 IRanges_1.21.23
[7] BiocGenerics_0.9.3
loaded via a namespace (and not attached):
[1] BatchJobs_1.1-1135 BBmisc_1.5 BiocParallel_0.5.8
bitops_1.0-6
[5] brew_1.0-6 codetools_0.2-8 DBI_0.2-7
digest_0.6.4
[9] fail_1.2 foreach_1.4.1 iterators_1.0.6 plyr_1.8
[13] RSQLite_0.11.4 sendmailR_1.1-2 stats4_3.1.0 tools_3.1.0
[17] zlibbioc_1.9.0
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--
Hervé Pagès
Program in Computational Biology
Division of Public Health Sciences
Fred Hutchinson Cancer Research Center
1100 Fairview Ave. N, M1-B514
P.O. Box 19024
Seattle, WA 98109-1024
E-mail: hpa...@fhcrc.org
Phone: (206) 667-5791
Fax: (206) 667-1319
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