Single-Indexed Sequencing Overview

The single-indexed sequencing workflow applies to all Illumina sequencing platforms, where an Index Read follows Read 1.

1.

Read 1—Read 1 follows the standard Read 1 sequencing protocol using SBS reagents. The Read 1 sequencing primer is annealed to the template strand during the cluster generation step.

2.

Index Read preparation—The Read 1 product is removed and the Index 1 (i7) sequencing primer is annealed to the same template strand. This step produces the Index 1 (i7) Read.

3.

Index 1 (i7) Read—Following Index Read preparation, the Index 1 (i7) Read is performed. The read length depends on the system and run parameters.

4.

Read 2 resynthesis—The Index Read product is removed and the original template strand is used to regenerate the complementary strand. Then, the original template strand is removed to allow hybridization of the Read 2 sequencing primer.

5.

Read 2—Read 2 follows the standard paired-end sequencing protocol using SBS reagents.