Insertion size when using 'paired' in MEDIPS.createSet()
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stb • 0
@stb-11175
Last seen 4.3 years ago

Hi, 

I have a question concerning insertion size when using MEDIPS.

When setting paired = TRUE in MEDIPS, you will get a mean insertion size as well as standard deviation;

Mean insertion size: 204.4319 nt
SD of the insertion size: 57.93392 nt
Max insertion size: 602 nt
Min insertion size: 49 nt

Does the insertion size include the reads or is it the number of nucleotides between the forward and reverse read?

Thanks, 

Stine

 

medips R medip-seq pair-end reads • 590 views
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Lukas Chavez ▴ 570
@lukas-chavez-5781
Last seen 3.7 years ago
USA/La Jolla/UCSD

Dear Stine,

the function getPairedGRange() contains the relevant interfaces to Rsamtools for importing bam files. These are:

(...)
scanFlag = scanBamFlag(isPaired = T, isProperPair = TRUE,
            hasUnmappedMate = FALSE, isUnmappedQuery = F, isFirstMateRead = T,
            isSecondMateRead = F, isSecondaryAlignment = isSecondaryAlignment)
(...)
scanParam = ScanBamParam(flag = scanFlag, simpleCigar = simpleCigar, what = c("rname",
                  "pos", "strand", "qwidth", "isize", "mpos”))
(...)
regions = scanBam(file = paste(path, fileName, sep = "/"),
            param = scanParam)
(…)

The insert size is accessed by
regions$isize

According to https://bioconductor.org/packages/devel/bioc/manuals/Rsamtools/man/Rsamtools.pdf, isize is defined as:
isize: This is the TLEN field in SAM Spec v1.4. Inferred insert size for paired end alignments.

The samtools definition on TLEN says:
If all segments are mapped to the same reference, the unsigned observed template length equals the number of bases from the leftmost mapped base to the rightmost mapped base.

Therefore, to the best of my knowledge, the insert size represents the entire sequenced DNA fragment.

Hope that helps.

All the best,
Lukas

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