Protocol

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This section describes the Illumina DNA Prep with Enrichment protocol.

• Review the planned complete sequencing workflow, from sample through analysis, to ensure compatibility of products and experiment parameters.
• Before proceeding, confirm kit contents and make sure that you have the required components, equipment, and consumables. This protocol requires library prep and enrichment reagents, an enrichment probe panel, and index adapters. Enrichment probe panels and index adapters are sold separately. Refer to Product Contents.
– Third-party biotinylated probes must meet specific requirements. Refer to Third-Party Panel Requirements to make sure that your third-party probes meet the requirements.
• Follow the protocol in the order shown, using the specified volumes and incubation parameters.

Illumina DNA Prep with Enrichment reagents are configured and tested at 1-plex and 12-plex enrichment plexity. Although other enrichment plexities are possible, some plexities require additional pre-enrichment library prep and enrichment probe panel reagents.

Obtaining suitable enrichment yields for nonstandard enrichment plexities might require additional optimization. Optimal results are not guaranteed. For information on reagents required for other enrichment plexities, refer to Non-Standard Enrichment Plexity Reagents.

• Enrichment plexity—The number of pre-enriched libraries (1–11 optional, 12 recommended) pooled together in one enrichment reaction for hybridization with the enrichment probe panels. For example, combining 12 pre-enriched libraries together creates a 12-plex enrichment pool.
• Enrichment reaction —The number of unique enrichment reaction preparations, regardless of the number of pre-enriched libraries pooled per reaction. For example, a single enrichment reaction can prepare a 1-plex or 12-plex enrichment pool.

To calculate the total number of post-enriched libraries, multiply the enrichment plexity per reaction by the number of enrichment reactions. For example, a single enrichment reaction of a 12-plex enrichment pool produces a pool of 12 post-enriched libraries.

When pooling pre-enriched libraries, Illumina DNA Prep with Enrichment reagents support the number of enrichment reactions and plexity indicated.

Supported Enrichment Plexities

Illumina DNA Prep with Enrichment Reagents

Enrichment Reactions

Enrichment Plexity

16-sample kit

16 reactions

1-plex

96-sample kit

8 reactions

12-plex

The following diagram illustrates this Illumina DNA Prep with Enrichment workflow.

• Safe stopping points are marked between steps
• Time estimates are based on processing 12 samples at 12-plex enrichment.