Installation

The Illumina NovaSeq 6000Dx Integration v1.3.0 supports the integration between Clarity LIMS and the NovaSeq 6000Dx instrument. The integration includes the following changes:

• Compatibility with Oracle Linux. For compatibility, refer to NovaSeq 6000Dx Integration v1.3.0 Release Notes.

• Security fixes. This document provides instructions for installing NovaSeq 6000Dx Integration v1.3.0 and describes the components that are installed in the default configuration.

If the user is new to the NovaSeq 6000Dx integration, then follow the instructions provided in this section. If the user has installed NovaSeq 6000Dx Integration v1.1.0, then note that there are no user-facing changes in the configured workflow between NovaSeq 6000Dx Integration v1.1.0 and NovaSeq 6000Dx Integration v1.3.0. It is safe to use the existing NovaSeq 6000Dx v1.1 workflow installed with IPP v2.5.0 and reconfigure the integration for this workflow instead. For more information, refer to [Optional] Configure Properties in Installation Steps.

Prerequisites

Prerequisite 1: Secret Utility Configuration

If Clarity LIMS Secret Utility is not installed or configured, use the following information to configure Secret Utility:

Installation on a hosted system with Clarity LIMS (v6.3 or later)

The Clarity LIMS installation tooling configures this installation. No additional configuration is necessary.

Installation on an on-premise Clarity LIMS instance (v6.3 or later)

The installation requires the following passwords:

• Tenant DB Password

• Clarity DB Password

• FTP password

• LDAP manager password (if LDAP is enabled)

• LabLink DB password

• Reporting DB password (if installed)

• RabbitMQ password

For more information on Secret Utility configuration, refer to the Clarity LIMS (Clarity & LabLink Reference Guide) documentation.

Prerequisite 2: Illumina Preset Protocols (IPP) v2.9

The Illumina NovaSeq 6000Dx Integration Package v1.3.0 depends on the configuration provided in Illumina Protocols (IPP) v2.9.

The installation procedure provided in this document includes steps that install the following IPP configurations on the server used for the NovaSeq 6000Dx integration installation:

• QC Protocols Base configuration

• NovaSeqDx configuration

• Library Prep Validation configuration (optional, but recommended)

For details on IPP v2.9 installation and configuration, refer to the Illumina Preset Protocols documentation.

Prerequisite 3: Proxy Update (Illumina Cloud Hosted Installation Only)

The Illumina cloud hosted installation configures an additional include directive for /etc/httpd/conf.d/clarity.conf in the proxy configuration. Additional proxy configuration files for the Sequencer API service are expected to be placed in the /etc/httpd/conf.d/clarity folder.

Sequencer API RPM Installation Components

NovaSeq 6000Dx Integration v1.3.0 is distributed as the BaseSpaceLIMS-sequencer-api RPM package. This package must be installed on the Clarity LIMS server.

The BaseSpaceLIMS-sequencer-api RPM installs the following files and scripts:

• Sequencer API WAR file

• application.yml configuration file

• configure_sequencer_api_proxy.sh configuration script

• configure_sequencer_api_env.sh configuration script

• configure_sequencer_api_application.sh configuration script

Sequencer API WAR

The RPM installs the Sequencer API WAR file into the same Tomcat container as Clarity LIMS. The WAR file is installed to the following location:

/opt/gls/clarity/tomcat/current/webapps/Illumina#Sequencer#v2##2.7.0.0.war

The version number can differ.

The WAR file is deployed automatically when Clarity LIMS launches. Because the RPM installs a new WAR file, Clarity LIMS is automatically stopped during installation of the RPM. Restart Clarity LIMS before installing the NovaSeq and Library Prep Validation workflow configuration from the IPP.

Configuration File

The application.yml configuration file is installed to the following location:

/opt/gls/clarity/extensions/sequencer-api/application.yml

The file properties are configured with a script during the installation process.

For a complete list of all properties included in the application.yml file and the properties installed into database, refer to Integration Properties Details.

Configuration Scripts

The scripts in the following table are installed to the following location:

/opt/gls/clarity/config

During installation, the RPM runs these scripts as they are needed.

Script Name	User	Description
configure_sequencer_api_proxy.sh	root	Adds configuration to the proxy to allow communication with the Sequencer API after Tomcat has been started. Run this script in the following scenarios: When the Sequencer API is first installed. When reconfiguring the Clarity LIMS proxy. This process happens until a version of Clarity LIMS that includes the necessary configuration files from /etc/httpd/clarity/*.conf is available. ℹ Not required for Illumina cloud hosted installations.
configure_sequencer_api_env.sh	root	Adds Secret Management setting into the Clarity LIMS Tomcat configuration. Run this script when the Sequencer API is first installed on a Secret Util-enabled Clarity LIMS.
configure_sequencer_api_application.sh	glsjboss	Configures the installed application.yml file. Sets the Clarity LIMS URL and the key to use when signing access tokens. ⚠ Run this script when the Sequencer API is first installed. If you run the script multiple times, the script reconfigures each setting to its default. The only exception is the signing key. For this setting, you are asked if you want a reset. If the reset is done, all previously issued tokens are no longer valid.

Installation Steps

If running the sequencing service and Automation Worker on the same instance, the instance must also be running a compatible version of the Oracle Linux operating system. For compatibility, refer to NovaSeq 6000Dx Integration v1.3.0 Release Notes.

[Optional] Install QC Protocols

If you are using PhiX control, install the base configuration of the QC Protocols from IPP as follows.

1. Log in as the root user on the server being used to install the integration.

2. Use the following yum command to install the IPP RPM:

   yum install BaseSpaceLIMS-Illumina-Preset-Protocols --enablerepo=<< repo name info from support >>

   The --enablerepo command-line argument must be included to enable the repo. The Illumina Support team can provide you with the repo file with the appropriate naming convention.

3. Enter y to confirm that you want to proceed with the RPM and package installation.

4. Make sure that NGS Extension Package v5.25.0 is installed.

   If this package is not installed, you must install it or upgrade the existing package.

5. As the glsjboss user, install the QC_Protocols.qc-protocols configuration from the following location:

   /opt/gls/clarity/config/illumina-preset-protocols-installer.sh -o install QC_Protocols.qc-protocols

For information on IPP installer commands, parameters, and options, refer to the Illumina Preset Protocols documentation.

Install Sequencer API

Install the Sequencer API as follows.

1. On the Clarity LIMS server, log in as the root user.

2. Use the following yum command to install the RPM:

   yum install BaseSpaceLIMS-sequencer-api --enablerepo=< repo name info from support >

   The --enablerepo command-line argument must be included to enable the repository. The Illumina Support team can provide you with the repo file with the appropriate naming convention.

3. Enter y to confirm that you want to proceed with the RPM installation.

4. If necessary, install the Secret Utility Package.

   For more information on this package, refer to Prerequisite 1: Secret Utility Configuration.
5. As the glsjboss user, run configure_sequencer_api_application.sh from the following location:

   bash /opt/gls/clarity/config/configure_sequencer_api_application.sh

6. Enter values for the presented as follows.

   You can also press Enter to accept the default values in the application.yml configuration file. For more information about the application.yml properties, refer to Integration Properties Details.

   • Would you like to use the apiuser user for communication with Clarity? [Y]

   • Enter required value for Token Expiry (hours) [88]

7. Update the Clarity LIMS proxy for the Sequencer API as follows.

   • For an on-premise system, run the following command:

     bash /opt/gls/clarity/config/configure_sequencer_api_proxy.sh

   • For an Illumina cloud hosted system, make sure that the proxy has been updated as follows. Place the sequencer-api.include file in /etc/httpd/conf.d/clarity.

     ProxyPass ajp://localhost:9009/Illumina/Sequencer/v2
     ProxyPassReverse ajp://localhost:9009/Illumina/Sequencer/v2

8. Update the Clarity LIMS Tomcat configuration as follows.

   1. Make sure that Secret Utility is installed and configured. For more information, refer to Prerequisite 1: Secret Utility Configuration.

   2. Run the following command to add the Secret Utility configuration to Clarity LIMS:

      bash /opt/gls/clarity/config/configure_sequencer_api_env.sh

      ℹ This script modifies /opt/gls/clarity/tomcat/current/bin/setenv.sh for Clarity LIMS Tomcat to load the Secret Utility environment variable file (/etc/profile.d/claritysecret.sh) during startup.

   3. Use the following command to Start Clarity LIMS:

      /opt/gls/clarity/bin/run_clarity.sh start

Install NovaSeqDx and Library Prep Validation Workflow Configuration

Install the NovaSeqDx and Library Prep workflow configuration from IPP as follows.

1. As the glsjboss user, run the following command to install the NovaSeq 6000Dx v1.2 workflow configuration:

   bash /opt/gls/clarity/config/illumina-preset-protocols-installer.sh -o install Illumina_Instruments.novaseqdx-v1.2

   ℹ The installer validates the import of the workflow and provides the Warning/Error details in STDOUT, which allows you to either proceed with the import or cancel it.

2. [Optional] Run the following command to install the Library Prep Validation v2.3.4 workflow:

   /opt/gls/clarity/config/illumina-preset-protocols-installer.sh -o install Illumina_Instruments.Library-Prep-Validation-v2.3

   This step is optional, but recommended. The Library Prep Validation v2.3.4 workflow allows you to validate the NovaSeqDx v1.2 workflow.

Check Sequencer API

Make sure that the Sequencer API is working as follows.

1. Visit https://<< Clarity LIMS URL >>/Illumina/Sequencer/v2/sequencing-run/login. This page returns the following URL string:

   https://<< Clarity LIMS URL >>/Illumina/Sequencer/v2/oauth/authorize?client_id=illumina_sequencer&redirect_uri=../sequencer_login&response_type=token

2. Copy and paste the URL string into the browser. The Clarity LIMS Sign In page opens.

3. Enter sign in credentials.

4. Select Sign In. A Successfully Authenticated message displays.

[Optional] Configure Properties

Configuring integration-related properties is optional.

For more information on the configurable integration properties that enable capture and generation of files associated with the sequencing run, refer to Integration Properties Details

Workflows, Protocols, and Steps Installed

NovaSeq 6000Dx Integration v1.3.0 includes the following workflows, protocols, and steps listed below. These workflows can be found in IPP v2.9.

Library Prep Validation v2.3.4 Workflow

Protocol: Library Prep Validation v2.3.4

Steps:

1. Library Prep Validation v2.3.4

NovaSeqDx v1.2 Workflow

Protocol 1: Run Format (NovaSeqDx v1.2)

Steps:

1. Define Run Format (NovaSeqDx v1.2)

Protocol 2: NovaSeqDx Standard (NovaSeqDx v1.2)

Steps:

1. Make Bulk Pool for NovaSeqDx Standard (NovaSeqDx v1.2)

2. Dilute and Denature (NovaSeqDx v1.2)

Protocol 3: NovaSeqDx Xp (NovaSeqDx v1.2)

Steps:

1. Make Bulk Pool for NovaSeqDx Xp (NovaSeqDx v1.2)

2. Dilute, Denature & ExAmp (NovaSeqDx v1.2)

3. Load to Flowcell (NovaSeqDx v1.2)

Protocol 4: AUTOMATED - NovaSeqDx Run (NovaSeqDx v1.2)

Steps:

1. AUTOMATED - NovaSeqDx Run (NovaSeqDx v1.2)

For detailed descriptions of the steps and automations included in each protocol, and details on other components in the configuration, refer to NovaSeq 6000Dx Integration v1.3.0 Configuration.

For instructions on using the Library Prep Validation v2.3.4 protocol, refer to NovaSeq 6000Dx Integration v1.3.0 User Interaction, Validation and Troubleshooting.

Instrument Software

The NovaSeq 6000Dx instrument software includes the following components:

• NovaSeq 6000Dx Operating Software (NVOS) — Contains the user interface for setting up the sequencing run. Responsible for controlling the instrument and acquiring the images.

• Real-Time Analysis v3 (RTA3) — Takes the images generated by the first module, processes, and analyzes them. Makes sure that data files are created and copied to the final destination folder.

• Sequencing Analysis Viewer (SAV) — Displays the important quality metrics generated by the RTA3 software.

• NovaSeq 6000Dx recipes — Provides system operation instructions for use with NovaSeq 6000Dx reagent kits for SP, S1, S2, and S4 flow cells.

• Universal Copy Service — Copies output files to destinations such as final destination folder and/or BSSH (when the instrument is configured for use with BSSH).

Instrument Integration Configuration

For the NovaSeq 6000Dx API integration to work, the instrument must be able to communicate with Clarity LIMS through the API. Complete the following steps to configure the NVOS and confirm that you can access Clarity LIMS from the instrument.

1. Launch NVOS and wait for the initialization process to complete.

2. On the NVOS Instrument Settings page, do as follows.

   1. Select the Global Settings tab.

   2. Under Instrument Mode, select one of the following modes:

      • Allow users to switch between IVD and RUO

      • IVD Only

      • RUO Only

      
3. If you selected IVD Only instrument mode, select the IVD Settings tab and complete the following steps:

   1. Configure Run Setup as follows.

      • Select the Select runs from the LIMS server option.

      • Enter the Clarity LIMS URL (e.g., https://yourserver.claritylims.com).

   2. [Optional] Under Output Location, enter the output folder location. You can also specify the output folder location when setting up a run in NVOS.

   
4. If you selected RUO Only instrument mode, select the RUO Settings tab and complete the following steps:

   1. Configure Run Setup as follows.

      • Select the Select runs from the LIMS server option.

      • Enter the Clarity LIMS URL(e.g., https://yourserver.claritylims.com).

   2. Configure Default Work Type as follows.

      • [Optional] If selection support between NovaSeq Xp or NovaSeq Standard is required for the workflow, select the Enable workflow selection checkbox.

      • Select the required default workflow type: NovaSeq Standard or NovaSeq Xp.

   3. [Optional] Configure Output Location as follows.

      • [Optional] Enter the output folder location. You can also specify the output folder location when setting up a run in NVOS.

      • [Optional] To upload and store run data in the cloud server, select the BSSH checkbox. Select a Configuration option (Run Monitoring and Storage or Run Monitoring Only) and select your Hosting Location from the drop-down list.

   
5. Select Save to complete the configuration.

6. On the NVOS home page, you can now select Sequence to log into Clarity LIMS and start a run. nova