What are primer pools? How many pools do I need?

We design panels based on our ability to multiplex tens of thousands of amplifications in a single tube. Amplicons that cannot be multiplexed in the same tube are put in separate tubes. Each separate tube of mixture of primers is called a primer pool.
In common cases, hotspot targets that are far apart from one another on the genome can be put in the same pool, no matter how many amplicons are involved; exon targets, or any region targets that are longer than the maximum allowed amplicon length, will need 2 pools to cover fully the region(s) of interest. This is because overlapping amplicons are not compatible in the same multiplexed reaction mix.
In very rare cases, a third or fourth pool, or even larger number of pools might be needed. Adding more pools beyond a 2-pool panel usually helps very little in increasing the in silico coverage of the targets, but might help in other aspects of the panel, such as adding the flexibility of splitting a larger panel into smaller panels post-ordering.

2018-09-12T13:56:50+00:00 September 12th, 2018|Comments Off on What are primer pools? How many pools do I need?