Bead size selection FAQ for Illumina DNA PCR Free

In the protocol, the IPB (Illumina Purification Beads) beads are added to BLT-PF beads—is this correct? Will the BLT-PF beads interfere with IPB purification? This reagent addition is correct; add IPB beads directly to library in the BLT-PF tube to improve efficiency and remove the necessity of an additional samples transfer step. The presence of BLT-PF beads does not interfere with size selection.

Can the incubation time for bead drying after size selection/ethanol wash be changed? Yes, if the bead pellets dry sooner, proceed with the elution. If after the standard dry time, there are any drops of liquid that were missed, remove them and dry again. Due to variations in environmental conditions, such as temperature and humidity, bead drying time may need to be extended or shortened.

What happens IPB beads are overdried? Over-drying of IPB beads will make final library elution less effective and may reduce final library yield.

What happens if some IPB beads are carried over in the samples? The final sample may not appear clear. In this case, final libraries containing residual IPB beads can be placed on a magnet for several minutes (until the liquid is clear and bead pellet has formed) and the library can be transferred to a new well or pooled for sequencing.

Is it possible to adjust library insert sizes? Illumina has tested IPB adjustments to obtain library sizes down to 350 bp and up to 550 bp fragments. For more details, see application note Tunable insert sizes with Illumina DNA PCR-Free Prep, Tagmentation.

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