How to plan a Local sequencing run on the NovaSeq X Series

Background

The NovaSeq X allows users to setup sequencing runs using Local or Cloud Run Planning, importing a Sample Sheet, or Manual mode. See the following steps to setup a sequencing run using Local Run Planning options.

For more information on Run Modes, refer to the Knowledge Base Article Run Mode options for the NovaSeq X Series

To setup a Manual Mode Run, refer to the Knowledge Base Article How to plan a Manual Mode NovaSeq X Series run for more information.

Create a Planned Run

1. Select the instrument icon at the bottom of the NovaSeq X Series Control Software to open the global navigation menu.

1. Select Runs, then navigate to the Planned tab and select Create Run.

1. In Run Settings, enter the applicable options:

• Run Name: A name to identify the current run (maximum of 255 alphanumeric characters, spaces, periods, dashes, and underscores are allowed).

• Run Description: [Optional and cannot contain asterisks, brackets, or commas].

• Generate FastQC metrics: If enabled, the analysis pipeline generates FastQC metrics.

• Read Lengths:

  • Read 1 — Enter the cycles number.

  • Index 1 — length of the index read for Index 1.

  • Index 2 — length of the index read for Index 2.

  • Read 2 — Enter the cycles number.

  • All Read Length fields must have a numerical value input. To skip a Read, input 0 for the number of cycles for that Read.

• Library tube ID: [Optional and is on the label of the library tube strip]. This setting can also be referred to as a sample container ID.

1. Select Next.

2. In Configuration, enter:

• Applications: Select the desired analysis applications, refer to Set Up Secondary Analysis.

• Description: [Optional]

• Library Prep Kit: Select a kit

  • When an Illumina library prep kit is selected, adapter sequences for Read 1 and Read 2 automatically populate and cannot be modified. Override cycles also automatically populate.

• Index Adapter Kit: Select a kit

  • For information on adding custom library prep and index adapter kits, refer to Import Custom Library Prep and Index Adapter Kits.

• Provide additional analysis settings as needed. The settings available or required vary based on the application, application version, and the configuration settings selected. For more information on DRAGEN secondary analysis, refer to Illumina DRAGEN Bio-IT Platform support page and the DRAGEN on NovaSeq X Series release notes that are available on the NovaSeq X Series support page.

1. Select Next.

2. Input the sample and indexes information (i5 indexes must be entered in the forward orientation).

• To import samples, select Download Template and fill out the Sample ID and Index information, then select Import Samples and import the filled-out template file.

1. Select Next.

2. Review the run configuration.

1. To enable multiple DRAGEN analysis configurations per run, select Add Another Configuration.

• NovaSeq X Control Software 1.3 and above: Up to 12 DRAGEN applications plus 1 BCL Convert application can be selected for both on-board and cloud analysis. Enter up to 32 Library Prep and Index Adapter kits combinations, per App (DRAGEN App + genome) configuration.

• Note:One sample can be loaded in multiple lanes (Sample 5 in Lane 1 and 2), but you cannot include the same sample for two different App Configuration.

1. Select Save As Planned to complete the run details and plan for sequencing.

Start a Planned Run

1. If you are not signed in, follow the instructions in Sign In and Sign Out.

2. Select Start.

3. Select Sequencing.

4. Select an instrument side for a single-sided run or select both sides for a dual-sided run.

• NovaSeq X Control Software version 1.4: If a single-sided run is in progress, the idle side of the instrument displays the next available time for a staggered start. For more information, refer to Staggered Start of Runs.

1. Select the current planned run.

Note: To change account or sign in the BaseSpace account select Change account button

1. Select Review to check the run configuration.

1. To use custom primers, select the appropriate Custom primer checkboxes.

• For more information on custom primers, refer to Custom Primers.

1. Confirm the output folder:

• To transfer run data to a local storage folder in addition to the cloud, provide an output folder.

• To only transfer run data to the cloud, select Don't transfer run data to external output folder.

• To change the default output folder, refer to Specify the Default Output Folder Location.

1. If performing local secondary analysis, confirm BCL data folder transfer setting.

• To update the default run data management selection, refer to Customize System Settings.

1. [Optional] Select a custom recipe file.

2. After reviewing the run information, select Load consumables.

After sequencing, Secondary Analysis starts with local DRAGEN on instrument.

For additional information refer to: NovaSeq X Plus Product Documentation, Plan a Sequencing Run

For any feedback or questions regarding this article (Illumina Knowledge Article #8528), contact Illumina Technical Support [e-mail].