can paired-end mates overlap?

[cmd]

Hi all,

I am pretty new to NGS and have a quick question. Can the two mates in a paired-end pair overlap each other, such as in the illustration below:

Insert: <---------------------------------------------------------->
Read1: <-------------------------------|......................................
Read2: ...................................|--------------------------------->

I have so far thought that could not be possible but I have seen some such instances from a dataset I am currently looking at.

Thanks

[kopi-o]

Yes, they can. This happens when the insert is shorter than (2*read length), as in your picture.

[boetsie]

As said; yes they can! Though i think your figure should be this for paired-ends (orientation issue, if you sequence with Illumina);

Fragment: <---------------------------------------------------------->
Read1 : ------------------------------->|......................................
Read2 : ..........................|<---------------------------------
Overlap: ..........................*****................................



The overlap depends on the fragment size and the number of cycles you sequence. If you have fragments of 250bp, you'll never get an overlap between the two reads, unless both reads are larger than 125bp (should be more to be able to find the actual overlap).

There are tools to do this too, see this thread;

http://seqanswers.com/forums/showthread.php?t=15742&highlight=FLASH

Regards,
Boetsie

[cmd]

Thank boetsie and kopi-o for your great answers

MD

[sphil]

keep in mind that some mappers handle those paired end like single end!

[pmiguel]

Okay, I think the nomenclature used here is confusing.

First, ignoring Roche terminology, "mate-pairs" refer to a library construction methodology that allows both ends of a large fragment of DNA to be captured in the same template. For next gen sequencers this involves circularizing the large fragment, destroying the non-circular DNA, then re-fragmenting the circles. The junction is somehow biotinylated, allowing fragments containing those junctions to be captured. Adapters are added to either end of this (smaller) fragment. Reads are obtained from either end.

Here is the confusion:

The OP, cmd, draws the orientation of reads against reference consistent with mate pair reads. However I do not believe it is possible for mate pair reads to overlap at their proximal (beginning) termini when mapped against a consensus.

If these are meant to be simple "paired end" reads then boetsie's diagram is correct and, yes PE reads can overlap in the middle.

Even that explanation ignores the two different meanings given to the term "mate pair" for 1st generation (sanger sequencing) and Roche GS-FLX reads, that are effectively "mate pair" but Roche calls "paired end"...

--
Phillip