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Obtaining a minimum threshold of 6 nM or 12 nM in a 20 µL volume after Normalase PCR, depending on the chosen workflow option, is required for expected functionality. Libraries that do not meet this threshold will be less than 2 nM or 4 nM post-Normalase Module (from 6 nM or 12 nM minimum thresholds, respectively) and will be under-represented during cluster generation.
Also, if multiple libraries are under 12 nM to start, the final library pool will be less than 4 nM, resulting in potential under-clustering of the pool on the flow cell. This is similarly the case for the 6 nM normalization workflow, where the final library pool will be less than 2 nM resulting in under-clustering of the pool on the flow cell.