low yield of SOLiD templated beads

[KevinLam]

Hi
We have been facing problems with solid 3 plus to come up with sufficient beads for deposition. We hope to sort this out before moving to solid 4.
I can't seem to find the troubleshooting manual for steps to tweak to increase yield. (I am new to the wet lab portion so please be gentle)

from the library prep stage What is the recommended template concentration?
I understand it's a balance betw having not enough template vs having more polyclonal beads

from the ePCR stage, sometimes we have to repeat the ePCR to increase the amt of templated beads to a level sufficient for deposition.
is there steps where we can increase the recovery efficiency? (maybe increase incubation times for some steps?)

[VanessaS]

Kevin,

For the 3+, we used .5 pM most of the time, with an occasional library at 1.0pM. We typically had about 68-75% monoclonal beads. A mini scale To get enough for a full slide, with really good deposition we had to do 4-5 ePCR reactions. One rxn is supposed to be enough for a quad.
If you are not already doing so, I highly recommend using qPCR with the taqman assay to get really accurate concentrations of your library prior to ePCR.