Configuration

This integration supports the integration of Clarity LIMS to Illumina MiSeq i100 series instruments.

The configuration provided in this integration has been established to support MiSeq i100 Series lab processes. Any configuration changes to protocols or workflows (including renaming protocols, steps, and fields) could break the integration.

Prerequisites and Assumptions

The MiSeq i100 Series Integration v1.1.0 requires a valid BaseSpace Sequence Hub account with an Enterprise or Professional subscription.

Analysis Configuration Template

The MiSeq i100 Series Sequencing v2.0 workflow uses analysis configuration templates (ACTs), created on BaseSpace Sequence Hub, to configure secondary analysis for planned runs.

The index adapter kit configured in the ACT must be created as label group in Clarity LIMS and used in the library preparation step for indexing the samples.

For more information, refer to MiSeq i100 Series Sequencing v2.0 Workflow User Interaction, Validation and Troubleshooting.

ACT names created in BaseSpace Sequence Hub must not have leading or trailing spaces.

Samples

Samples must go through the library preparation and quantification process before they enter the MiSeq i100 Series Sequencing v2.0 workflow. It is assumed that the following steps have completed before samples are assigned to the workflow:

• Samples have been accessioned into Clarity LIMS.

• Samples have been run through QC and library prep.

• Samples have the Molarity (nM) global field set to a value.

  This value is required for the Calculate Volumes automation in the Make Bulk Pool step.

Sample and library names must use only alphanumeric, dash, or underscore characters.

For more information on sample accessioning, refer to Sample Accessioning and Upload and Modify Samples in the Getting Started section of the Clarity LIMS (Clarity & LabLink Reference Guide) documentation.

You can assign samples to workflows automatically, using a routing script, or manually—from the Projects & Samples dashboard. Refer to Assign and Process Samples in the Clarity LIMS (Clarity & LabLink Reference Guide) documentation.

Workflows, Protocols, and Steps

The MiSeq i100 Series Integration Package v1.1.0 includes the following workflows:

• MiSeq i100 Series Sequencing v2.0

• Library Prep Validation v2.3.6 (optional, but recommended for validation purposes)

The following describes the protocols and steps included in these workflows.

Library Prep Validation v2.3.6 Workflow

Protocol 1: Library Prep Validation v2.3.6

Purpose:

• Included for validation purposes only, this protocol models the library prep steps required to advance samples to the MiSeq i100 Series Sequencing v2.0 workflow.

  ⚠ The label group (index adapter kit) used for library preparation must be the same as the index adapter kit specified in the ACT that is being used. For more information, refer to MiSeq i100 Series Sequencing v2.0 Workflow User Interaction, Validation and Troubleshooting.

• The protocol contains a single step - Library Prep Validation v2.3.6. After this step, a routing script sends the samples to the first step of the MiSeq i100 Series Sequencing v2.0 workflow (Assign Analysis Configuration Template (MiSeq i100 Series Sequencing v2.0)).

Steps:

1. Library Prep Validation v2.3.6

MiSeq i100 Series Sequencing v2.0 Workflow

Protocol 1: MiSeq i100 Series Sequencing v2.0

Purpose:

• This protocol models the lab processes of run setup for starting a MiSeq i100 Series sequencing and secondary analysis run.

Steps:

1. Assign Analysis Configuration Template (MiSeq i100 Series Sequencing v2.0)

   • The secondary analysis of samples is configured using the ACT.

   • The labels applied to the samples are validated against the selected ACT.

2. Make Bulk Pool (MiSeq i100 Series Sequencing v2.0)

   • The Calculate Volume script assists with pooling samples and adding resuspension buffers and reagents.

   • The Validate Analysis Configurations automation validates the ACT of pooled samples to make sure that the secondary analysis configurations remain valid after pooling.

3. Load To Dry Cartridge (MiSeq i100 Series Sequencing v2.0)

   • The library pool from step 2 is ready to be loaded to the MiSeq i100 Series dry cartridge.

   • Validates run and analysis configuration.

   • Generates the sample sheet and creates the planned run, depending on the run mode.

4. AUTOMATED - Sequencing Run (MiSeq i100 Series Sequencing v2.0)

   • The step is fully automated and starts and completes automatically after the sequencing run is started and completed in BaseSpace Sequence Hub.

   • The sequencing run metadata (e.g., run configuration, primary run metrics, etc.) are recorded automatically.

5. AUTOMATED - Analysis Run (MiSeq i100 Series Sequencing v2.0)

   • The step is fully automated and starts and completes automatically after the analysis run is started and completed in ICA.

   • The analysis run metadata (e.g., status and demultiplexing stats file) are recorded automatically.

Validation Workflow

The Library Prep Validation v2.3.6 workflow allows for validation of the system after installation is complete. This workflow can be replaced by other custom library preparation workflows.

Configure a custom library preparation workflow as follows.

1. Select Configuration, then select Lab Work.

2. Add a Routing Script automation to the last Master Step of the library preparation workflow.

   
3. Select Configuration, then select Automation.

4. Create a new Routing Script automation and update it as follows.

   1. For Channel Name, enter limsserver.

   2. For Command Line, enter the following script:

      bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -u {username} -p {password} -i {stepURI:v2} -l {compoundOutputFileLuid0} script:changeWorkflow \
      \
      --FIELD_NAME 'Sequencing Instrument' \
      --FIELD_VALUE 'MiSeq i100 Series' \
      --WORKFLOW 'MiSeq i100 Series Sequencing v2.0' \
      --STEP 'Assign Analysis Configuration Template (MiSeq i100 Series Sequencing v2.0)' \
      --INPUTS_OR_OUTPUTS 'OUTPUTS'"

   3. For Automation Use, select the custom library preparation workflow.

   

How the Integration Works

The following information summarizes how the MiSeq i100 Series integration works.

• After the Validate Run Setup and Create Planned Run automation is triggered on the Load to Dry Cartridge (MiSeq i100 Series Sequencing v2.0) step, validation of the run and analysis configuration occurs. When the validation fails, Clarity LIMS displays an error message. Otherwise, a cloud-planned run is created and available on BaseSpace Sequence Hub. The v2 sample sheet is also generated.

  • The planned run contains the run and analysis configuration information required to start the run on the instrument. The sample sheet is generated and stored in Clarity LIMS for reference purposes.

  • In Cloud run mode, analysis is performed in Illumina Connected Analytics (ICA). In Hybrid run mode, analysis is performed on-premise.

• When the integration is configured with BaseSpace Sequence Hub storage and monitoring, the integration tracks the sequencing and analysis events. You can view the analysis result files in BaseSpace Sequence Hub.

If a planned run with the same sample name and project name (case-insensitive) has been created previously, the sample sheet generated from the Validate Run Setup and Create Planned Run automation can reflect the original case of the previous sample name. This can cause validation errors for analysis configurations with sample-level settings. To resolve this issue, change the sample name or the project name on Clarity LIMS and run the automation again.

Start a Sequencing Run on MiSeq i100 Series Instrument

The following sections outline the steps to start a Clarity LIMS cloud or hybrid run on the MiSeq i100 Series instrument.

Run Mode Configuration

1. In the MiSeq i100 Series Control Software, log in to the instrument using an account that has permissions for configuring settings.

2. On the menu, select Settings, then select Cloud settings.

3. In Run Settings, select the Cloud run storage checkbox.

4. For Hosting location, select the applicable location from the drop-down options.

5. [Optional] For Private domain name, enter the domain name according to the workgroup that was configured with the MiSeq i100 Series integration package.

6. Select Save.

Start a Planned Run (Cloud Run or Hybrid Run Modes)

Refer to information on starting a planned run in the Illumina support site..

Enabling Planned Run Generation for Samples Having Duplicate Name with Different Indexes

The library preparation workflow of the samples must be configured to ensure unique derived sample names before routing the samples through the library preparation workflow.

Components Installed

The following sections describe the components (files, properties, reagent categories / label groups, reagent kits, and containers) that are installed by default as part of this integration.

Reagent Kits

• MiSeq i100 Series Dry Cartridge

• MiSeq i100 Series Library Denature (KLD)

• MiSeq i100 Series Wet Cartridge

• Resuspension Buffer (RSB)

Container Types

• MiSeq i100 Series Dry Cartridge

• Tube

Rules and Constraints

• The workflow configuration contains several validation checks. To make sure that the calculations work properly, it is important that you do not disable any of this validation logic. Refer to the individual validation checks for details.

• For all submitted samples that require analysis, they must have an associated secondary analysis that is configured using the analysis configuration template (ACT). The ACT must be configured on BaseSpace Sequence Hub before starting the Assign Analysis Configuration Template step. The ACT names must be unique.

• The dry cartridge barcode must be unique. There must not be multiple dry cartridges with the same name in the system.

• Reagent labels, or indexes, must be unique in a single sequencing run.

• Only one library pool can be added to be Load to Dry Cartridge step.

• The AUTOMATED - Sequencing Run (MiSeq i100 Series Sequencing v2.0) and AUTOMATED - Analysis Run (MiSeq i100 Series Sequencing v2.0) steps must not be manually started or completed. These steps are fully automated and the sequencing service does not update samples correctly if they have been manually started.

• For the automated run to start successfully, you must select Validate Run Setup and Create Planned Run in the Load to Dry Cartridge step. The Load to Dry Cartridge step must be completed and libraries must be queued to the AUTOMATED - Sequencing Run step.