The Illumina MiSeq instrument integration provides preconfigured workflows that map to established lab protocols and steps used with the MiSeq instrument. The integration also supports the associated library prep kits, reagent kits, and assays used in these protocols.

The Illumina MiSeq Integration v8.3.0 includes the following features:

• Preconfigured MiSeq Sequencing v3.2 workflow that maps to lab protocol steps and instrument runs.

• Automated sample sheet generation for use with MiSeq Control Software (MCS) and Local Run Manager.

• The following supported Local Run Manager analysis modules:

  • Generate FASTQ

  • Library QC

• Automated tracking of the sequencing run and the parsing of the following run statistics to Clarity LIMS:

  • Sequencing run metrics

  • Sequencing run parameters

• Preconfigured Library Prep Validation workflow that is used for validation purposes only. This workflow contains a single step protocol that models the library prep required to produce normalized libraries that are ready for the MiSeq Sequencing v3.2 workflow. MiSeq Integration Package v8.3.0 works with Library Prep Validation v2.3.4, which is installed with Illumina Preset Protocols (IPP) v2.9. For details, refer to

MiSeq Integration v8.3.0 runs as independent service and can be on the same server as other Clarity LIMS integrations. The integration is only required for MCS v4.0. If you are running MCS v3.0 or below, do not upgrade to this integration.

Last Updated: July 2025

Release Date: July 2024

Document Version: 3

These release notes describe the key changes to software components for the Clarity LIMS MiSeq Integration Package v8.3.0.

Compatibility

Last Updated: July 2025

Release Date: September 2023

Document Version: 3

These release notes describe the key changes to software components for the Clarity LIMS MiSeq Integration Package v8.2.0.

Compatibility

The Illumina MiSeq Integration v8.2.0 includes the following features:

• Preconfigured MiSeq Sequencing v3.2 workflow that maps to lab protocol steps and instrument runs.

• Automated sample sheet generation for use with MiSeq Control Software (MCS) and Local Run Manager.

• The following supported Local Run Manager analysis modules:

  • Generate FASTQ

  • Library QC

• Automated tracking of the sequencing run and the parsing of the following run statistics to Clarity LIMS:

  • Sequencing run metrics

  • Sequencing run parameters

• Preconfigured Library Prep Validation workflow that is used for validation purposes only. This workflow contains a single step protocol that models the library prep required to produce normalized libraries that are ready for the MiSeq Sequencing v3.2 workflow. MiSeq Integration Package v8.2.0 works with Library Prep Validation v2.3.1, which is installed with Illumina Preset Protocols (IPP) v2.6. For details, refer to

MiSeq Integration v8.2.0 runs as independent service and can be on the same server as other Clarity LIMS integrations. The integration is only required for MCS v4.0. If you are running MCS v3.0 or below, do not upgrade to this integration.

MiSeq Integration v8.2.0 includes the MiSeq v3.2 workflow upgrade. This upgrade is typically done through the API. To upgrade the workflow manually, create the Validate Single Input automation and update its trigger information as follows.

Create Validate Single Input Automation

1. In Clarity LIMS, under Configuration, select the Automation tab.

2. Select New Automation.

3. Create an automation called Validate Single Input.

Change Trigger Point for the Validate Single Input Automation

1. In Clarity LIMS, under Configuration, select the Lab Work tab.

2. Find the Denature and Dilute step.

3. In the Automation section, update the Trigger Location and Trigger Style for the Validate Single Input automation as follows.

The Illumina MiSeq Integration Package v8.3.0 supports the integration of Clarity LIMS to MiSeq instruments.

MiSeq Integration v8.3.0 runs as independent service and can be on the same server as other Clarity LIMS integrations. The integration is only required for MCS v4.0. If you are running MCS v3.0 or below, do not upgrade to this integration.

Compatibility

MiSeq Integration v8.3.0 is compatible with the following software:

• Clarity LIMS v6.2.0 and later

• Secret Util v1.0 and later

• IPP v2.9 and later

• MCS v4.0/LRM v3.0 and later

• Oracle Linux (for compatibility, refer to )

Prerequisites

MiSeq Integration v8.3.0 has the following prerequisites:

• Mount run data network-attached storage (NAS) share

• IPP is installed

Installation

MiSeq Integration v8.3.0 is distributed as the ClarityLIMS-Illumina-MiSeq-Package-v8 RPM package that must be installed on the Clarity LIMS server. This package installs the following items:

• Bash scripts used to run the service

• The miseq-sequencing.jar file

• The configure_extensions_miseq_sequencingservice.sh script

The following installation steps are required for the installation of MiSeq Integration v8.3.0.

Install the RPM

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

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

   ℹ You must use the --enablerepo command line argument to enable the repo. For the repo file and the correct name to use, contact Illumina Support.

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

Configure the Integration Properties

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

Start the Sequencing Service

Run the following command to start the sequencing service:

Workflows, Protocols, and Steps Installed

MiSeq Integration v8.3.0 works with the MiSeq Sequencing v3.2, which contains a single protocol with the same name. This protocol includes the following steps:

• Library Pooling (MiSeq v3.2)

• Denature and Dilute (MiSeq v3.2)

• MiSeq Run (MiSeq v3.2)

For descriptions of the protocol and the steps, refer to . For instructions on using the Library Prep Validation v2.3.4 workflow to validate the automated sample sheet generation, refer to .

Instrument Software

The MiSeq Control Software (MCS) is divided into the following modules:

• MCS — Controls the instrument operation, including various configuration settings. This software is installed and runs on the instrument.

• Local Run Manager — Analyzes sequencing data and integrates with MCS. This software can be accessed on the instrument through a web browser.

• Real-Time Analysis (RTA) — Performs image processing and base calling (primary analysis). The software makes sure that data files are created and copied to the final destination folder and is installed and runs on the instrument.

For more information on MCS, refer to the MiSeq documentation at .

Instrument Integration

The instrument integration must be performed and maintained by Illumina Support. Illumina Support requires remote access to the instrument while it is idle.

The support team has created batch files that use custom scripts during the key events of a sequencing run. When these batch files are used, they read the event information and write it in a TXT event file at the same network share location that the instrument uses to write the run data. Another process running on the Clarity LIMS server receives the event files and takes the appropriate actions.

The sequencing service monitors the end of the run event. This event is used to capture key step data and files and parse run statistics for output custom fields.

Validation

The following sections explain how to validate the installation of the Illumina MiSeq Integration Package v8.3.0.

The validation involves the following processes:

• Running samples through the Library Prep Validation v2.3.4 workflow. The workflow contains a single-step protocol that models the library prep required to produce normalized libraries. At the end of the step, the normalized libraries automatically advance to the MiSeq Sequencing v3.2 workflow.

• Running normalized libraries through the MiSeq Sequencing v3.2 workflow. This process validates the following information:

The following sections explain how to validate the installation of the Illumina MiSeq Integration Package v8.2.0.

The validation involves the following processes:

• Running samples through the Library Prep Validation v2.3.1 workflow. The workflow contains a single-step protocol that models the library prep required to produce normalized libraries. At the end of the step, the normalized libraries automatically advance to the MiSeq Sequencing v3.2 workflow.

• Running normalized libraries through the MiSeq Sequencing v3.2 workflow. This process validates the following information:

  • Automated generation of a sample sheet for use with the MiSeq Control Software (MCS) and Local Run Manager (LRM).

  • Automated tracking of the MiSeq sequencing run and parsing of run statistics into Clarity LIMS, including the following information:

    • Run status and metrics of sequencing run

Activate Workflow, Create Project, Add and Assign Samples

The following steps prepare Clarity LIMS to run samples through the Library Prep Validation and MiSeq Sequencing v3.2 workflows.

1. In the Clarity LIMS Configuration area, select each workflow and change the Workflow Status toggle to Active.

2. On the Projects & Samples screen, create a project. Add samples to the project.

3. Assign the samples to the Library Prep Validation workflow.

Library Prep Validation Protocol

This single-step protocol models the library prep required to produce normalized libraries that are ready for the MiSeq Sequencing v3.2 workflow.

Follow the steps in to run the Library Prep Validation workflow with the following:

• Label Group = TruSeq HT Adapters v2 (D7-D5)

• Sequencing Instrument = MiSeq

On exit from the step, the Routing Script automation is triggered. This automation assigns samples to the first step of the MiSeq Sequencing v3.2 workflow, which is Library Pooling (MiSeq v3.2) step.

MiSeq Sequencing v3.2 Protocol

Run Library Pooling (MiSeq v3.2)

1. In Lab View, locate the MiSeq Sequencing v3.2 protocol. Samples are queued for the Library Pooling (MiSeq v3.2) step. Select the step to proceed to the Queue screen.

2. On the Queue screen, do as follows.

   1. Add the samples to the Ice Bucket.

Run Denature and Dilute (MiSeq v3.2)

1. In Lab View, locate the MiSeq Sequencing v3.2 protocol. The pooled samples are queued for the Denature and Dilute (MiSeq v3.2) step. Select the step to proceed to the Queue screen.

2. On the Queue screen, add the pool to the Ice Bucket, and then select View Ice Bucket.

   ⚠ Only one pool is allowed as an input.

Run MiSeq Run (MiSeq v3.2)

1. In Lab View, locate the MiSeq Sequencing v3.2 protocol. The pool of samples is queued for the MiSeq Run (MiSeq v3.2) step. Select the step to proceed to the Queue screen.

2. On the Queue screen, add the pool to the Ice Bucket, and then select View Ice Bucket.

3. On the Ice Bucket screen, select Begin Work.

Using Step Details Presets

Most of the custom fields in Step Details are analysis settings used by Local Run Manager (LRM) and are for advanced users only. The following fields are exceptions:

• Workflow

• Experiment Name

• Description

• Read 1 Cycles

To use the default analysis settings, select the analysis module from the Preset drop-down list.

The default values populate the respective fields and indicate the following actions are needed:

• Required — The field is required for the specific analysis module selected. Replace the word Required with a proper value. For example, the Genome Folder field should contain the directory to the genome folder.

• None — The field is not applicable for the specific analysis module selected. Leave the field as is.

For a list of fields that are applicable for the analysis module selected, refer to the Applicable analysis fields for the selected Workflow field in Step Details.

Validating Creation of Event Files

This validation checks the following information:

• The destination path is correctly configured.

• The instrument computer can access and write to the destination path.

• There are no syntax errors in the Clarity LIMS batch file.

Use the following steps to confirm that event files are created by the batch file in the destination path. The steps assume that the default configuration has been successfully imported.

1. In C:\Illumina\gls, double-click the batch file gls_event_mcs_rta_lrm.bat.

2. Confirm that an empty event file appears in the configured DESTINATION_PATH.

Manual execution of the batch file produces an output resembling the following example:

Troubleshooting

If an automation trigger does not appear to run the corresponding scripts, see the following topics:

• Troubleshooting Automation Worker in the .

• Troubleshooting Automation in the .

If an error occurs that does not provide direction on how to proceed, confirm the version of the installed MiSeq Integration Package. To confirm the version, run the

command from the server console.

If the error is related to data parsing, retrieving run results data, or report values not appearing as expected, review the MiseqIntegrator.log file. The file is located at

Additional troubleshooting information for this integration is provided in the Illumina Instrument Integrations FAQ.

If you are unable to resolve the issue, contact the Clarity LIMS support team. Supply the relevant information from the troubleshooting already performed.

The Illumina MiSeq Integration Package v8.2.0 supports the integration of Clarity LIMS to MiSeq instruments.

MiSeq Integration v8.2.0 runs as independent service and can be on the same server as other Clarity LIMS integrations. The integration is only required for MCS v4.0. If you are running MCS v3.0 or below, do not upgrade to this integration.

Compatibility

MiSeq Integration v8.2.0 is compatible with the following software:

• Clarity LIMS v6.2.0 and later

• Secret Util v1.0 and later

• IPP v2.6 and later

• MCS v4.0/LRM v3.0 and later

• Oracle Linux (for compatibility, refer to )

Prerequisites

MiSeq Integration v8.2.0 has the following prerequisites:

• Mount run data network-attached storage (NAS) share

• IPP is installed

Installation

MiSeq Integration v8.2.0 is distributed as the ClarityLIMS-Illumina-MiSeq-Package-v8 RPM package that must be installed on the Clarity LIMS server. This package installs the following items:

• Bash scripts used to run the service

• The miseq-sequencing.jar file

• The configure_extensions_miseq_sequencingservice.sh script

The following installation steps are required for the installation of MiSeq Integration v8.2.0.

Install the RPM

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

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

   ℹ You must use the --enablerepo command line argument to enable the repo. For the repo file and the correct name to use, contact Illumina Support.

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

Configure the Database Service Properties

To enable capture and generation of files associated with the sequencing run, configure the following properties using the omxProps-ConfigTool utility.

Configurable Database Properties

Start the Sequencing Service

Run the following command to start the sequencing service:

Workflows, Protocols, and Steps Installed

MiSeq Integration v8.2.0 works with the MiSeq Sequencing v3.2, which contains a single protocol with the same name. This protocol includes the following steps:

• Library Pooling (MiSeq v3.2)

• Denature and Dilute (MiSeq v3.2)

• MiSeq Run (MiSeq v3.2)

For descriptions of the protocol and the steps, refer to . For instructions on using the Library Prep Validation v2.3.1 workflow to validate the automated sample sheet generation, refer to .

Instrument Software

The MiSeq Control Software (MCS) is divided into the following modules:

• MCS — Controls the instrument operation, including various configuration settings. This software is installed and runs on the instrument.

• Local Run Manager — Analyzes sequencing data and integrates with MCS. This software can be accessed on the instrument through a web browser.

• Real-Time Analysis (RTA) — Performs image processing and base calling (primary analysis). The software makes sure that data files are created and copied to the final destination folder and is installed and runs on the instrument.

For more information on MCS, refer to the MiSeq documentation at .

Instrument Integration

The instrument integration must be performed and maintained by Illumina Support. Illumina Support requires remote access to the instrument while it is idle.

The support team has created batch files that use custom scripts during the key events of a sequencing run. When these batch files are used, they read the event information and write it in a TXT event file at the same network share location that the instrument uses to write the run data. Another process running on the Clarity LIMS server receives the event files and takes the appropriate actions.

The sequencing service monitors the end of the run event. This event is used to capture key step data and files and parse run statistics for output custom fields.

Validation

The Illumina MiSeq Integration Package v8.3.0 supports the integration of Clarity LIMS to Illumina MiSeq Sequencing Systems.

The integration allows for automated tracking of an Illumina sequencing run in Clarity LIMS. This capability includes tracking sequencing run status, generating run report, and capturing and parsing run statistics. In addition, this integration provides automated generation of a sample sheet file for use with the MiSeq Control Software (MCS) and Local Run Manager (LRM).

For instructions on user interaction for each step, validating and troubleshooting the Illumina MiSeq Integration Package, refer to MiSeq Integration v8.3.0 User Interaction, Validation and Troubleshooting.

Compatibility

• The MiSeq Sequencing v3.2 workflow is compatible with MiSeq Integration Package v8.2.0 and v8.3.0.

• MiSeq Integration v8.2.0 and above is only required for MiSeq Control Software (MCS) v4.0. For MCS v3.1 or earlier, do not upgrade to MiSeq Integration v8.2.0 or above. This upgrade breaks the integration.

Prerequisites and Assumptions

Before samples are assigned to the MiSeq Sequencing v3.2 workflow, make sure that the following prerequisites are completed:

• Samples have been accessioned into Clarity LIMS.

• Samples have been run through QC and library prep.

• Samples have been normalized, and the value is captured in a field called Normalized Molarity (nM).

For more information on sample accessioning, refer to Sample Accessioning and Upload and Modify Samples in the .

Samples can be assigned to the MiSeq Sequencing v3.2 workflow automatically using a routing script or manually from the Projects & Samples dashboard. Refer to Assign and Process Samples in the .

Workflows, Protocols, and Steps

The Illumina MiSeq Integration includes the MiSeq Sequencing v3.2 workflow, which contains a single protocol of the same name.

Step 1: Library Pooling (MiSeq v3.2)

In this step, the lab scientist manually places libraries into pools in the Clarity LIMS Placement screen.

Master Step Fields

The following field is configured on the Library Pooling (MiSeq v3.2) master step. The field displays on the Record Details screen at run time.

Library Pooling (MiSeq v3.2) Master Step Field Configuration

Global Fields

The following table lists the global fields that are displayed on the Queue and Ice Bucket screens of the Library Pooling (MiSeq v3.2) step. Most fields display in expanded view only.

Global Field Configuration (Submitted Sample)

Global Field Configuration (Derived Sample)

Step 2: Denature and Dilute (MiSeq v3.2)

In this step, pooled libraries are denatured and diluted, and then placed into the reagent cartridge loaded into the MiSeq instrument.

Master Step Fields

Most fields configured on the Denature and Dilute (MiSeq v3.2) step display on the Record Details screen in the Step Data table.

These fields are manually populated at run time. The values are then used to generate the sample sheet.

Denature & Dilute (MiSeq v3.2) Master Step Field Configuration

Groups of Defaults

Global Fields

The following table shows the global fields that are configured to display on the Queue, Ice Bucket, and Record Details screens of the Denature and Dilute (MiSeq v3.2) step:

Global Field Configuration (Submitted Sample)

Global Field Configuration (Derived Sample)

Step File Placeholders

Placeholders for the following files are configured on the Record Details screen of the Denature and Dilute (MiSeq v3.2) step.

Step 3: MiSeq Run (MiSeq v3.2)

Master Step Fields

The following fields are configured on the MiSeq Run (MiSeq v3.2) step. These fields display on the Record Details screen at run time. The read-only fields are automatically populated at the end of the run.

MiSeq Run (MiSeq v3.2) Master Step Field Configuration

Global Fields

There are several sample and measurement global fields that are displayed on the Record Details screen of the MiSeq Run (MiSeq v3.2) step. These fields are autopopulated at the end of the sequencing run.

For more information, refer to .

Step File Placeholders

Placeholders for the following files are configured on the Record Details screen of the MiSeq Run (MiSeq v3.2) step:

• Illumina Run Report (automatically attached)

• Link to Run Folder (automatically attached)

• Run Parameters (automatically attached)

For details, refer to .

Sample Sheet Generation

Sample sheet generation occurs in the Denature & Dilute (MiSeq v3.2) step. This step places samples on the container loaded in the system.

The default configuration provides only the Validate Run Setup and Generate MiSeq SampleSheet automation. This automation uses the Template File Generator (DriverFileGenerator.jar) and a template file to generate a CSV format file for use with the MiSeq Control Software (MCS).

The sample sheet content is determined by the fields on the Record Details screen of the step in the Step Data table. The values entered into these fields are used to populate the sample sheet.

To customize the template used to create the sample sheet, you can insert additional columns.

The following additional details are available:

• For a sample template that you can download and customize for the lab, refer to .

• For instructions on customizing the template, refer to .

Sequencing Results Parsing

The MiSeq Run (MiSeq v3.2) step records information for the flow cell lane and generates a report summarizing the results. In addition, run parameters, run info, and a link to the run folder are automatically captured.

Generated and Captured Files

The following table describes the run information files, reports, placeholders, and links that Clarity LIMS automatically generates or captures during a sequencing run:

Run Information Generated or Captured by MiSeq Run (MiSeq v3.2) Step

Metadata

The following list shows metadata that Clarity LIMS automatically captures from the Illumina sequencing software as part of a sequencing run. This information is gathered from various run result files and events.

• Chemistry

• Experiment Name (entered in software)

• Finish Date (run completion date)

Primary Analysis Metrics

The following table lists the real-time analysis (RTA) primary analysis metrics Clarity LIMS automatically captures and records per read, for samples in each flow cell lane. These metrics are captured upon run completion and are stored as fields in the Sample Details table on the Record Details screen.

To see both per read and per lane metrics, expand the output.

RTA Primary Analysis Metrics Captured by MiSeq Run (MiSeq v3.2) Step

How It Works

1. The sequencing service runs on the Clarity LIMS server. The service detects event files that instrument RTA produces as the run progresses. The event files let the service know where to find the run data.

2. As the run data are written out and the End Run event is detected, the following events occur:

   • The data are matched to the step based on the reagent cartridge ID that was entered or scanned on the Denature and Dilute (MiSeq v3.2) step.

Scripts and Files Installed

This integration requires components installed with the Illumina Preset Protocols (IPP).

The Illumina MiSeq Integration Package v8.3.0 RPM installs the scripts and files listed in the following table.

Properties Installed

Refer to for the properties installed with Illumina MiSeq Integration v8.3.0.

Consumables Installed

Reagent Categories and Kits

Reagent categories or label groups are installed with the IPP workflow configuration slices.

The MiSeq Reagent Kit is included in the Illumina MiSeq Integration.

Control Types

The PhiX v3 control type is included in the Illumina MiSeq Integration.

Container Types

The following container types are included in the Illumina MiSeq Integration:

• MiSeq Reagent Cartridge

• 96 well plate

• Tube

All one-dimensional container types with both numeric rows and numeric columns are supported.

Instrument Integration

To make sure that the Illumina instrument warranty remains valid, the instrument integration must be performed and maintained by the Clarity LIMS Support team. To perform this integration, the Support team requires remote access to the system while it is idle.

The following steps are performed by the Clarity LIMS Support team when configuring the sequencing for use with the Illumina MiSeq Integration.

1. Create a directory on the local computer to hold the batch files. These batch files write event files to the network attached storage (NAS) shares.

2. Create a directory on the NAS to hold the event files.

3. Modify Illumina software configuration files to call the batch files that create the event files.

Rules and Constraints

The Illumina MiSeq Integration operates with the following constraints:

• The reagent cartridge ID must be unique. There should not be multiple reagent cartridge containers in the system with the same name.

• The reagent cartridge ID must be scanned as the reagent cartridge Container Name on the Denature & Dilute (MiSeq v3.2) step.