Custom Primers

Follow the run setup instructions described in the Run Settings. Using custom primers for a run on an Illumina iSeq 100 Sequencing System requires three additional steps at the run setup:

• Set the iSeq 100 to manual run mode and enable the custom primers setting.
• Select the custom primers in the control software during run setup.
• Prepare and add the appropriate volume of each custom primer to the library reservoir of the cartridge after pre-run checks are completed.

To use custom primers with the iSeq 100 i1 Reagent v2 kits, update and install iSeq 100 Control Software v3.0 or later. The iSeq 100 Sequencing System support pages on the Illumina support center provides additional resources.

The iSeq 100 allows the use of two custom read primers and two custom index primer combinations. Each primer is loaded into the library reservoir individually.

Depending on your library preparation kit, you might be required to use Illumina primer mixes. Refer to VP10 and VP14 Custom Primers for more information.

Illumina cannot guarantee the performance or compatibility of custom primers. You are responsible for validating custom primers for sequencing on the iSeq 100 Sequencing System.

To enable custom primers , update and install iSeq 100 Control Software v3.0 or later. The iSeq 100 must be set to manual run mode. Local Run Manager is not compatible with custom primers. Refer to Set Up a Sequencing Run (Manual Mode) for more information.

1. In the control software menu, select System Settings.
2. Select the Customization tab.
3. Select Enable custom primers.
4. Select Save, then select Exit.

When custom primers are selected during run setup, the software directs the system to pull reagents from the library reservoir. Illumina primers are not used for the read or index during the sequencing run. Illumina primers refer to primers already in the reagent cartridge wells.

If Illumina primers are not used for Read 1 or Read 2, the optional PhiX control is not sequenced.

Although the PhiX is not indexed, it does provide sequencing data during the index read. Sequencing data from the PhiX control is not generated during the index read, regardless of which indexing primer is used.