Technical data

Post-Capture Sample Processing for Multiplexed Sequencing 5
Step 6. Optional: Pool captured libraries for sequencing
SureSelect
XT2
Target Enrichment System for Illumina 73
Step 6. Optional: Pool captured libraries for sequencing
See Table 32 on page 74 for post-capture pooling guidelines, based on
your SureSelect
XT2
Capture Library size and sequencing design. Pooling
instructions are provided below.
1 Combine the capture pools such that each index-tagged sample is
present in equimolar amounts in the final sequencing sample pool. For
each final pool, use the formula below to determine the amount of each
capture pool to use.
Volume of capture pool
Vf Cf
# Ci
---------------------------------
= where
where V(f) is the final desired volume of the sequencing sample pool,
C(f) is the desired final concentration of all the DNA in the pool
# is the number of capture pool samples to be combined, and
C(i) is the initial concentration of each capture pool sample.
2 Adjust the final volume of the pooled library to the desired final
concentration.
• If the final volume of the combined index-tagged samples is less than
the desired final volume, V(f), add Low TE to bring the volume to
the desired level.
• If the final volume of the combined index-tagged samples is greater
than the final desired volume, V(f), lyophilize and reconstitute to the
desired volume.
Table 32 shows an example of the amount of 2 capture pool samples (of
different concentrations) and Low TE needed for a final volume of
20 µL at 10 nM final DNA concentration.