Insert size for paired end sequencing for identification of structural variation
Dear all,
Does anyone have any experience of the ideal fragment size for paired end sequencing looking at structural variation in whole genome sequencing?
I am doing a pilot project to whole genome sequence a single individual per lane. It may be the best strategy to choose a small size of 200bp and then run the same sample again using mate pair sequencing and much larger insert size of 2kb. This way we would achieve the optimum trade off between detection and resolution of break points, as outlined in the following paper;
Designing deep sequencing experiments: detecting structural variation and estimating transcript abundance.
Bashir A, Bansal V, Bafna V.
Just doing paired end 75bp sequencing. Using the GAII the insert size can be 200-500bp. depending on how well the sequencing works and funding etc I may not get to do the mate pair part, so will just end up with the shorter fragment paired end data. With this in mind it might be better to go larger than 200bp and aim for 500bp. Does anyone have any experience of using this fragment size? It is ideal to aim for below 500bp, as this is the maximum length for cluster generation. If there are some reads over 500bp I guess they would reduce cluster efficiency?
Any comments on this would be really appreciated.
Thanks!
Does anyone have any experience of the ideal fragment size for paired end sequencing looking at structural variation in whole genome sequencing?
I am doing a pilot project to whole genome sequence a single individual per lane. It may be the best strategy to choose a small size of 200bp and then run the same sample again using mate pair sequencing and much larger insert size of 2kb. This way we would achieve the optimum trade off between detection and resolution of break points, as outlined in the following paper;
Designing deep sequencing experiments: detecting structural variation and estimating transcript abundance.
Bashir A, Bansal V, Bafna V.
Just doing paired end 75bp sequencing. Using the GAII the insert size can be 200-500bp. depending on how well the sequencing works and funding etc I may not get to do the mate pair part, so will just end up with the shorter fragment paired end data. With this in mind it might be better to go larger than 200bp and aim for 500bp. Does anyone have any experience of using this fragment size? It is ideal to aim for below 500bp, as this is the maximum length for cluster generation. If there are some reads over 500bp I guess they would reduce cluster efficiency?
Any comments on this would be really appreciated.
Thanks!