Bead clean up ratios used in the Illumina 16S demonstrated protocol
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Below are the bead clean-up ratios used after each PCR step of the Illumina 16S protocol.
PCR Clean‐Up 1
This step uses purification beads to purify the 16S V3 and V4 amplicon away from free primers and primer dimer species.
25 µl of samples + 20 µl beads = 0.8X beads:DNA ratio
PCR Clean‐Up 2
This step uses purification beads to clean up the final library before quantification.
50 µl of libraries + 56 µl beads = 1.12X beads:DNA ratio
Note: As the beads:DNA ratio changes, the length of fragments binding to the beads and left in solution also changes--the lower the ratio of beads:DNA, the larger the final fragments will be at elution.
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