Bead clean-up ratios used in the Illumina 16S Metagenomic Sequencing Library Preparation protocol

Below are the bead clean-up ratios used after each PCR step of the 16S Metagenomic Sequencing Library Preparation demonstrated 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-to-sample 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-to-sample ratio

Note: As the beads-to-sample ratio changes, the length of fragments binding to the beads and left in solution also changes; the lower the ratio of beads-to-sample, the larger the final fragments will be at elution.

For any feedback or questions regarding this article (Illumina Knowledge Article #6127), contact Illumina Technical Support [e-mail].