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When doing Illumina 2x150bp sequencing of genomic DNA, and after aligning the reads to GRCh38, does the percentage of the non-N fraction of the human genome as MAPQ=0 depend on the insert sizes of the genomic fragments?

This is, for two identical samples with identical final coverage, sample A having an average insert size of 250bp and sample B having an average of 450bp, would the fraction of MAPQ=0 change between the two?

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Yes, as a general rule of thumb mappability increases with insert size (up to a limit) and read length. Whether this will actually occur in a given case will depend more on how randomly the sequencing samples from the genome to begin with (i.e., if the library prep happens to select for/against high mappability regions then the insert size won't much matter). The basic reason behind this is that if one end of a read aligns to a repetitive element that the likelihood of being able to use the other end as an anchor will increase a bit if it's further away. Now having said that, Illumina sequencers in particular like a certain fragment length for efficient sequencing, so you can't go too crazy on increasing the insert sizes and still get decent output (unless you modify your library prep).

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This is, for two identical samples with identical final coverage, sample A having an average insert size of 250bp and sample B having an average of 450bp, would the fraction of MAPQ=0 change between the two?

For human, I expect sample B will have fewer MAPQ=0 reads (I don't have concrete data to prove). A large part of human genome consists of ALU repeats. The average size of an ALU is ~320bp. If your insert size is ~450bp, you can map reads into most of isolated ALUs in principle. With ~250bp insert, fragments in the middle of ALUs won't be mappable uniquely.

Note that to see the difference, you need to use a mapper that sufficiently takes the advantage of paired-end information.

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  • $\begingroup$ Does bwamem take advantage of it? $\endgroup$
    – 719016
    Jan 11 '18 at 8:18

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