The Illumina NovaSeq X Series Integration Package v1.2.0 supports the integration of Clarity LIMS to Illumina NovaSeq X series instruments.
For instructions on validating and troubleshooting the Illumina NovaSeq X Series Integration, refer to NovaSeq X Series Integration v1.2.0 User Interaction, Validation and Troubleshooting.
The configuration provided in this integration has been established to support NovaSeq X Series lab processes. Any configuration changes to protocols or workflows (including renaming protocols, steps, and fields) could break the process.
The NovaSeq X Series Integration v1.2.0 requires a valid BaseSpace Sequence Hub account with an Enterprise or Professional subscription.
The NovaSeq X Series Sequencing v1.1 workflow uses analysis configuration templates (ACTs) to configure secondary analysis for planned runs. Required ACTs must be created using BaseSpace Sequence Hub. The ACT names must be unique. The index adapter kit (the label group in Clarity LIMS) selected in the ACT is created in Clarity LIMS. The same label group must be used in the library preparation step. For more information, refer to NovaSeq X Series Integration v1.2.0 User Interaction, Validation and Troubleshooting.
Make sure that ACT names created in BaseSpace Sequence Hub do not have leading or trailing spaces. Otherwise, Clarity LIMS can have issues with recognizing the ACT names.
Samples must go through the library preparation and quantification process before they enter the NovaSeq X Series Sequencing v1.1 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.
The NovaSeq X Series Integration Package v1.2 includes the following workflows:
NovaSeq X Series Sequencing v1.1
Library Prep Validation v2.3.2 (optional, but recommended for validation purposes)
The following describes the protocols and steps included in these workflows.
The Library Prep Validation v2.3.2 workflow allows for validation of the system after installation is complete. This workflow can be replaced by other custom library preparation workflows. For details, refer to NovaSeq X Series Integration v1.2.0 User Interaction, Validation and Troubleshooting.
Configure a custom library preparation workflow as follows.
Select Configuration, then select Lab Work.
Add a Routing Script automation to the last Master Step of the library preparation workflow.
Select Configuration, then select Automation.
Create a new Routing Script automation and update it as follows.
For Channel Name, enter limsserver.
For Command Line, enter the following script:
For Automation Use, select the custom library preparation workflow.
The Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1) step uses ACTs to configure secondary analysis for samples. Each ACT contains details related to the secondary analysis (e.g., index adapter kit to use, reference genome setting, and so on). You can select an ACT from a list and assign samples to it. A planned run can have multiple ACTs and this step must be repeated for each ACT that is required for the run. For more information on creating ACTs and assigning samples to them, refer to NovaSeq X Series Integration v1.2.0 User Interaction, Validation and Troubleshooting.
¹ These automations are required for the NovaSeq X Series Sequencing v1.1 workflows and contain additional logic needed for CLPA support. If you would like to remove support for CLPA, contact Illumina Support.
² These automations are required for CLPA support only.
All remaining automations are required for the NovaSeq X Series Sequencing v1.1 workflows.
The following table lists the configuration details for fields that are defined on the Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1) step.
Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1) Master Step Field Configuration
The following table lists the global custom fields that are configured to display on the Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1) step.
Global Field Configuration (Derived Sample)
Samples are pooled in the Make Bulk Pool (NovaSeq X Series Sequencing v1.1) step. You can manually create working pools based on the final loading concentration required.
¹These automations are required for CLPA support only.
All remaining automations are required for the NovaSeq X Series Sequencing v1.1 workflows.
The following table lists configuration details for the fields that are defined on the Make Bulk Pool (NovaSeq X Series Sequencing v1.1) step.
Make Bulk Pool (NovaSeq X Series Sequencing v1.1) Master Step Field Configuration
The following table lists the global custom fields that are configured to display on the Make Bulk Pool (NovaSeq X Series Sequencing v1.1) step.
Global Custom Fields Configuration (Derived Sample)
The Dilute and Denature (NovaSeq X Series Sequencing v1.1) step allows you to dilute pooled samples with the addition of RSB.
¹ These automations are required for the NovaSeq X Series Sequencing v1.1 workflows and contain additional logic needed for CLPA support. If you would like to remove support for CLPA, contact Illumina Support.
² These automations are required for CLPA support only.
All remaining automations are required for the NovaSeq X Series Sequencing v1.1 workflows.
The following table lists the global custom fields that are configured to display on the Dilute and Denature (NovaSeq X Series Sequencing v1.1) step.
Global Field Configuration (Derived Sample)
The Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) step allows you to scan the library tube strip barcode into Clarity LIMS. Then, you can place the working pools into the library tube strip used in the NovaSeq X Series run. This step also does the following actions:
Validates the run setup and analysis information.
Generates the sample sheet file.
Creates a planned run on Illumina Connected Analytics (ICA), depending on the selected run mode.
¹ These automations are required for the NovaSeq X Series Sequencing v1.0 workflows and contain additional logic needed for CLPA support. If you would like to remove support for CLPA, contact Illumina Support.
² These automations are required for CLPA support only.
All remaining automations are required for the NovaSeq X Series Sequencing v1.0 workflows.
The following table shows the master step fields that are configured on the Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) step. These fields are required for sample sheet generation and planned run creation in ICA.
Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) Master Step Field Configuration
The AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step is fully automated. The integration starts the step, adds samples to the Ice Bucket, and completes the step. Data from the run is parsed back to Clarity LIMS. In this step, pooled samples in the library tube strip are sequenced on the NovaSeq X Series instrument.
¹ These automations are required for CLPA support only.
The following tables show the master step fields that are configured on the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step.
Clarity LIMS Master Step Field Information
Other Clarity LIMS Master Step Fields
The following global custom fields are used to capture the run metrics in Clarity LIMS:
% Bases >=Q30 R1
% Bases >=Q30 R2
% Error Rate R1
% Error Rate R2
Yield (Gb) R1
Yield (Gb) R2
Reads PF R1
Reads PF R2
%PF R1
%PF R2
% Aligned R1
% Aligned R2
% Phasing R1
% Phasing R2
% Prephasing R1
% Prephasing R2
Intensity Cycle 1 R1
Intensity Cycle 1 R2
Cluster Density R1
Cluster Density R2
At the end of the step, the pools of samples are automatically removed from the step. The step completes automatically when Run Status is RunCompletedSuccessfully.
The AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) step is fully automated. The integration starts and completes the step automatically. Data from the analysis is parsed back to Clarity LIMS. In this step, the secondary analysis configured using the Analysis Configuration Template (ACT) is performed in Illumina Connected Analytics (ICA). There are no automations associated with this step.
The following tables show the master step fields that are configured on the AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) step.
AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) Master Step Field Information
The following information summarizes how the NovaSeq X Series integration works.
After the Validate Run Setup and Create Planned Run automation is triggered on the Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) step, the run parameters entered in the Run Details screen are sent to Illumina Connected Analytics (ICA). The analysis configuration parameters from the selected ACTs are also sent to ICA. ICA validates the run and analysis configuration. If the validation fails, ICA sends an error message to Clarity LIMS. If the validation passes, ICA generates the sample sheet and sends it back to Clarity LIMS. ICA also creates the planned run based on the selected run mode (e.g., Local or Cloud).
For the Local run mode, the sample sheet is generated and stored in Clarity LIMS. This sample sheet contains the run and analysis configuration information required to start the run on the NovaSeq X Series instrument.
For the Cloud run mode, the planned run is created in ICA. 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.
When the sequencing run starts on the instrument, the NovaSeq X Series Control Software notifies BaseSpace Sequence Hub. BaseSpace Sequence Hub monitors the run and notifies Clarity LIMS via the integration service. The events are processed and the integration service retrieves the run information from BaseSpace Sequence Hub. This information is used to populate the custom fields in the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step.
Other run events follow the same information flow. When sequencing is complete, the control software uploads the sequencing run data (primary metrics) and associated files to ICA. Then, Clarity LIMS retrieves the primary metrics and uses them to populate the fields in the Sample Details table (e.g., % Error Rate R1). The custom fields (e.g., Run Status, Current Read, and so on) on the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step are updated using the run related information. If the sequencing run is successfully completed, the step automatically completes.
The integration tracks the analysis events and results in the AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) step. You must log in to BaseSpace Sequence Hub to retrieve the cloud analysis results. For local analysis, the secondary analysis results are in the external storage configured in Illumina Run Manager. The external storage information is found in the External Storage for Analysis Results configuration settings in Illumina Run Manager.
The following sections outline the steps to start a Clarity LIMS cloud or local run on the NovaSeq X Series instrument.
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.
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
Buffer Cartridge
Lyophilization Cartridge
NaOH
Reagent Cartridge
Resuspension Buffer (RSB)
TT2
Container Types
Library 2-Tube Strip
Library 8-Tube Strip
Tube
This integration supports the library 8-tube strip with the barcode in the LC[0-9]{7}-L[A-Z]1 format. The integration also supports the library 2-tube strip with the barcode in the LC[0-9]{7}-L[A-Z]2 format.
Control Types
PhiX v3
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. The validation checks determine the following information:
Which samples, and how many, can enter each step together.
Which samples, and how many, can be pooled together.
All submitted samples 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 library tube strip barcode must be unique. There must not be multiple library tube strip containers with the same name in the system.
Reagent labels, or indexes, must be unique.
One library pool can only contain one library or control with no label/index.
The AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) and AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) 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 Library Tube step.
NovaSeq X Series Integration Package v1.2.0 includes modifications to the existing NovaSeq X Series Sequencing workflows to support the following new features:
PhiX control included in the Dilute and Denature step instead of the Make Bulk Pool step.
Support for the new 1.5B and 25B flow cells.
Analysis tracking for the status and high level analysis result summary through the AUTOMATED - Analysis Run step (this is done at the metaworkflow level).
Analysis tracking is a new step in NovaSeq X Series protocol and cannot be added to a non-pending protocol or workflow. For analysis tracking at the metaworkflow level, do not proceed with the manual upgrade. Instead, install the NovaSeq X Series v1.2.0 Integration Package with the NovaSeq X Series Sequencing v1.1 protocol.
The following workflow updates are included in NovaSeq X Series Integration Package v1.2.0:
AUTOMATED - Sequencing Run step: Renamed the Library Tube Barcode master step custom field. This field is now named Library Tube Strip Barcode.
Load to Library Tube Strip step: Removed the Output Folder master step custom field.
Make Bulk Pool master step: Updated the dropdown items in the Final Loading Concentration (pM) custom field.
Use the following instructions to manually update the workflows.
Illumina recommends upgrading to Illumina Preset Protocols (IPP) v2.7 to support the new features. If you do not have IPP v2.7 installed, do the following before starting the manual upgrade:
Install ClarityLIMS-NGS-Package-v5 RPM v5.24.
Replace the following files in /opt/gls/clarity/extensions/conf/driverfiletemplates:
NovaSeqXSeries_Bulk_Pool1.csv:
NovaSeqXSeries_Bulk_Pool2.csv:
NovaSeqXSeries_Dilute_Denature_Calculate_Volumes.csv:
On the Configuration tab, select Consumables, and then select Containers.
Add the following information to enable the Library 2-tube Strip container that supports the 1.5B flow cell:
For Container Name, enter Library 2-tube Strip.
For Rows, update the following fields:
Number: 2
Naming: Alphabetic
For Columns, update the following fields:
Number: 1
Naming: Numeric
Start at: 1
Enable PhiX control in the Dilute and Denature step as follows.
On the Configuration tab, select Custom Fields, and then select Master Step Fields.
Add the following master step fields for the Dilute and Denature (NovaSeq X Series Sequencing v1.x) step:
Dilute and Denature (NovaSeq X Series Sequencing v1.x) Master Step Field Configuration
On the Configuration tab, select Custom Fields, and then select Master Step Fields.
Modify the following master step fields:
Make Bulk Pool (NovaSeq X Series Sequencing v1.x) Master Step Field Modifications
Load to Library Tube Strip (NovaSeq X Series Sequencing v1.x) Master Step Field Modifications
AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.x) Master Step Field Modifications
On the Configuration tab, select Custom Fields, and then select Global Fields.
For the NovaSeq X Flowcell Type field, update the Dropdown Items with the following values:
1.5B
10B
25B
Add Automation
On the Configuration tab, select Automation, and then select Step Automation.
On the Load to Library Tube Strip (NovaSeq X Series Sequencing v1.x) step, add the following automation to enable new flow cell types:
Name: Validate Flowcell Inputs and Validate Analysis Configurations and Register Step Started
Channel Name: limsserver
Command line:
Configure Automation Settings
On the Configuration tab, select Lab Work.
Configure the trigger location and style for the following automations:
Load to Library Tube Strip (NovaSeq X Series Sequencing v1.x) Step Automation Configuration
Dilute and Denature (NovaSeq X Series Sequencing v1.x) Step Automation Configuration
Modify Step Automations
On the Configuration tab, select Automation, and then select Step Automation.
Select the Calculate Volumes automation that is enabled on the Make Bulk Pool (NovaSeq X Series Sequencing v1.x) step.
Update the existing command line as follows.
This modification is required to move the PhiX control from the Make Bulk Pool (NovaSeq X Series Sequencing v1.x) step to the Dilute and Denature (NovaSeq X Series Sequencing v1.x) step. Changes are in red.
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} -log {compoundOutputFileLuid1} \ script:evaluateDynamicExpression \ -exp 'if (!input.hasValue(::Molarity (nM)::)) { return; }; if (output.hasValue(::Number of Samples in Pool::)) { output.::Number of Samples in Pool:: = output.::Number of Samples in Pool:: + 1; } else { output.::Number of Samples in Pool:: = 1; }' -t true \ script:evaluateDynamicExpression \ -exp 'output.::Bulk Pool Volume (ul):: = 100 * step.::Number of Lanes to Sequence::; output.::NovaSeq X Flowcell Type:: = step.::Flowcell Type::; output.::Final Loading Concentration (pM):: = step.::Final Loading Concentration (pM)::; input.::Per Sample Volume (ul):: = 2 * output.::Bulk Pool Volume (ul):: / input.::Molarity (nM):: / output.::Number of Samples in Pool::; output.::Total Sample Volume (ul):: = 0;' -t true \ script:calculate_multipool_adjusted_per_sample_volume -t \ script:evaluateDynamicExpression \ -exp 'if (output.hasValue(::Total Sample Volume (ul)::)) { output.::Total Sample Volume (ul):: = output.::Total Sample Volume (ul):: + input.::Adjusted Per Sample Volume (ul)::; } else { output.::Total Sample Volume (ul):: = input.::Adjusted Per Sample Volume (ul)::; }' -t true \ script:evaluateDynamicExpression \ -exp 'if (output.::Total Sample Volume (ul):: > output.::Bulk Pool Volume (ul)::) { output.::RSB Volume (ul):: = 0 } else { output.::RSB Volume (ul):: = output.::Bulk Pool Volume (ul):: - output.::Total Sample Volume (ul):: }' -t true \ && /opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/DriverFileGenerator.jar -i {stepURI:v2} -u {username} -p {password} -l {compoundOutputFileLuid1} script:driver_file_generator \ -t /opt/gls/clarity/extensions/conf/driverfiletemplates/NovaSeqXSeries_Bulk_Pool1.csv -o 1.csv \ script:driver_file_generator \ -t /opt/gls/clarity/extensions/conf/driverfiletemplates/NovaSeqXSeries_Bulk_Pool2.csv -o 2.csv \ && cat 1.csv 2.csv > {compoundOutputFileLuid0}.csv \ && /opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} -log {compoundOutputFileLuid1} \ script:evaluateDynamicExpression \ -exp 'output.::Number of Samples in Pool:: = ::::; output.::Total Sample Volume (ul):: = ::::; output.::Bulk Pool Volume (ul):: = ::::;' -t true \ && echo 'Calculate Volume completed successfully'"
Select the Calculate Volumes automation that is enabled on the Dilute and Denature (NovaSeq X Series Sequencing v1.x) step.
Update the existing command line as follows.
This modification is required to move the PhiX control from the Make Bulk Pool (NovaSeq X Series Sequencing v1.x) step to the Dilute and Denature (NovaSeq X Series Sequencing v1.x) step and to enable support for the 1.5B and 25B flow cells. Changes are in red.
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} -log {compoundOutputFileLuid1} \ script:evaluateDynamicExpression \ -exp ' if (input.::NovaSeq X Flowcell Type:: == ::10B:: || input.::NovaSeq X Flowcell Type:: == ::1.5B::) { output.::NaOH Volume (ul):: = 8.5; output.::TT2 Volume (ul):: = 127.5; output.::BP Aliquot Volume (ul):: = input.::Final Loading Concentration (pM):: * 170 / (2 * 1000); output.::RSB Volume (ul):: = 34 - output.::BP Aliquot Volume (ul)::; if (step.::1-2% PhiX Spike-In::) { output.::PhiX Volume (ul):: = 1; output.::PhiX Concentration (pM):: = 300; } else { output.::PhiX Volume (ul):: = ::::; output.::PhiX Concentration (pM):: = ::::; }; } else { output.::NaOH Volume (ul):: = 14; output.::TT2 Volume (ul):: = 210; output.::BP Aliquot Volume (ul):: = input.::Final Loading Concentration (pM):: * 280 / (2 * 1000); output.::RSB Volume (ul):: = 56 - output.::BP Aliquot Volume (ul)::; if (step.::1-2% PhiX Spike-In::) { output.::PhiX Volume (ul):: = 1.6; output.::PhiX Concentration (pM):: = 300; } else { output.::PhiX Volume (ul):: = ::::; output.::PhiX Concentration (pM):: = ::::; }; }' && /opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/DriverFileGenerator.jar -i {stepURI:v2} -u {username} -p {password} \ script:driver_file_generator \ -t /opt/gls/clarity/extensions/conf/driverfiletemplates/NovaSeqXSeries_Dilute_Denature_Calculate_Volumes.csv -o {compoundOutputFileLuid0}.csv -q true -destLIMSID {compoundOutputFileLuid0} -l {compoundOutputFileLuid1} \ && echo; echo 'Calculate Volumes completed successfully.'"
Select the Set Next Step automation that is enabled on the Dilute and Denature (NovaSeq X Series Sequencing v1.x) step.
Update the existing command line as follows.
This modification is required to enable support for the 1.5B and 25B flow cells and future 8-tube library strips. Changes are in red.
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} script:evaluateDynamicExpression -t false -h false -exp 'output.::Run Mode:: = input.::Run Mode::; output.::NovaSeq X Flowcell Type:: = input.::NovaSeq X Flowcell Type::; nextStep = ::ADVANCE::' -log {compoundOutputFileLuid1}"
Select the Validate Library Tube Strip Barcode automation that is enabled on the Load to Library Tube Strip Barcode (NovaSeq X Series Sequencing v1.x) step.
Update the existing command line as follows.
This modification is required to enable support for the 1.5B and 25B flow cells and future 8-tube library strips. Changes are in red.
bash -l -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -i {stepURI:v2} -u {username} -p {password} script:validate_output_containers -l {compoundOutputFileLuid1} -r 'Library 8-tube Strip:LC[0-9]{7}-L[A-Z]{1}1' -r 'Library 2-tube Strip:LC[0-9]{7}-L[A-Z]{1}2' -max 1"
Enable PhiX Control
On the Configuration tab, select Lab Work.
Select the Make Bulk Pool (NovaSeq X Series Sequencing v1.x) step.
For Control Types, remove PhiX v3.
For the Record Details milestone, navigate to Step Data and remove the following fields:
Final Loading Volume (ul)
% PhiX (2.0 nM) Spike-In
Select the Dilute and Denature (NovaSeq X Series v1.x) step.
Select the Record Details milestone.
Navigate to Step Data and add 1-2% PhiX Spike-In to Master Step Fields.
Enable New Flow Cell Types
On the Configuration tab, select Lab Work.
Select the Load to Library Tube Strip (NovaSeq X Series Sequencing v1.x) step.
Select the Queue milestone.
Navigate to Sample Table and add the NovaSeqX Flowcell Type field to Column Headers.
Select the Ice Bucket milestone.
Navigate to Sample Table and add the NovaSeqX Flowcell Type field to Column Headers.
Select the Placement milestone.
Navigate to Destination Containers and add Library 2-tube Strip.
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 NovaSeq X Series Integration v1.2.0 User Interaction, Validation and Troubleshooting.
Do not add samples to the Ice Bucket or start or complete the step. The integration does this automatically.
Do not add samples to the Ice Bucket or start or complete the step. The integration does this automatically.
This script is required for the NovaSeq X Series Sequencing v1.1 workflow to function properly.
The BP Aliquot Volume value is rounded up to the nearest µl. The value in the calculation file, however, is reflected with up to two decimal places.
This script is required for the NovaSeq X Series Sequencing v1.1 workflow to function properly.
Do not add samples to the Ice Bucket or start and complete the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step. The integration does this automatically.
Do not add samples to the Ice Bucket or start and complete the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step. The integration does this automatically.
Do not modify the names of the master step fields. Modifications cause the integration to break.
If a planned run with the same sample name and project name (case-insensitive) has been created previously in ICA, 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.
No action is required for the 25B flow cell. The flow cell uses the same Library 8-tube Strip as the 10B flow cell.
A merge conflict is expected when upgrading from IPP v2.6 to v2.7 due to the changes in the NovaSeq X Flowcell Type field.
These configuration updates are required to enable the new flow cell types.
This configuration update is required to enable PhiX control on the Make Bulk Pool (NovaSeq X Series Sequencing v1.x) step.
Field Name
Field Type
Options
Analysis Configuration Template
Text Dropdown
Required Field
Application
Text
Read Only
Application Version
Text
Read Only
Index Adapter Kit
Text
Read Only
Library Prep Kit
Text
Read Only
Reference Genome
Text
Read Only
Secondary Analysis Mode
Text
Read Only
Field Name
Field Type
Options
Additional Options and Dropdown Items
Molarity (nM)
Numeric
Decimal places displayed = 2
ACT Name
Text
Read Only
Field Name
Field Type
Options
Additional Options and Dropdown Items
Final Loading Concentration (pM)
Numeric Dropdown
Required Field
Custom Entries
Dropdown Items
90
140
150
160
180
Range from 0
Decimal places displayed: 0
Flowcell Type
Text
Required Field
Dropdown Items
1.5B
10B
25B
Minimum Per Sample Volume (ul)
Numeric
Required Field
Decimal places displayed: 2
Default: 2
Number of Lanes to Sequence
Numeric
Required Field
Range: 1–10
Decimal places displayed: 0
Field Name
Field Type
Options
Additional Options and Dropdown Items
Final Loading Concentration (pM)
Numeric Dropdown
Required Field
Custom Entries
Decimal places displayed: 0
Dropdown Items
225
400
RSB Volume (ul)
Numeric
Read Only
Decimal places displayed: 2
NovaSeq X Flowcell Type
Text Dropdown
Required Field
Read Only
Dropdown Items
1.5B
10B
25B
Field Name
Field Type
Options
Additional Options and Dropdown Items
BP Aliquot Volume (ul)
Numeric
Read Only
Decimal places displayed = 0
NaOH Volume (ul)
Numeric
Read Only
Decimal places displayed = 2
RSB Volume (ul)
Numeric
Read Only
Decimal places displayed = 2
TT2 Volume (ul)
Numeric
Read Only
Decimal places displayed = 2
Field Name
Source File
Corresponding Field in Source File
Additional Information
Current Cycle
RunCompletionStatus.xml
CompletedReads
Visible on Record Details screen.
Calculated based on the CompletedReads field.
Current Read
RunCompletionStatus.xml
RunParameters.xml
CompletedReads
PlannedReads
Visible on Record Details screen.
Calculated based on the CompletedReads field in RunCompletionStatus.xml and PlannedReads field in RunParameters.xml.
Flow Cell Expiration Date
RunParameters.xml
ExpirationDate
Visible on Record Details screen.
Corresponds to the value in the ExpirationDate field under the ConsumableInfo field for FlowCell Type.
Flow Cell ID
RunParameters.xml
SerialNumber
Visible on Record Details screen.
Corresponds to the value in the SerialNumber field under the ConsumableInfo field for FlowCell Type.
Flow Cell Lot Number
RunParameters.xml
LotNumber
Visible on Record Details screen.
Corresponds to the value in the LotNumber field under the ConsumableInfo field for FlowCell Type.
Flow Cell Part Number
RunParameters.xml
PartNumber
Visible on Record Details screen.
Corresponds to the value of the PartNumber field under the ConsumableInfo field for FlowCell Type.
Flow Cell Side
RunParameters.xml
Side
Visible on Record Details screen.
Flow Cell Type
RunParameters.xml
FlowCellType
Visible on Record Details screen.
Instrument Control Software Version
RunParameters.xml
SystemSuiteVersion
Visible on Record Details screen.
Instrument ID
RunParameters.xml
InstrumentSerialNumber
Visible on Record Details screen.
Instrument Type
RunParameters.xml
InstrumentType
Visible on Record Details screen.
Output Folder
RunParameters.xml
OutputFolder
Visible on Record Details screen.
Library Tube Strip Barcode
RunParameters.xml
SerialNumber
Visible on Record Details screen.
Corresponds to the value of the SerialNumber field under the ConsumableInfo field for SampleTube Type.
Library Tube Lot Number
RunParameters.xml
LotNumber
Hidden
Corresponds to the value of the LotNumber field under the ConsumableInfo field for SampleTube Type.
Run End Time
RunCompletionStatus.xml
RunEndTime
Hidden
Run Name
RunParameters.xml
ExperimentName
Visible on Record Details screen.
RTA Version
Not used
Not used
Hidden
Field Name
Description
Additional Information
Run Status
Presets
RunStarted
RunCompletedSuccessfully
RunAbortedByUser
RunErroredOut
Set by the integration service.
Visible on Record Details screen.
Sequencing Log
Set by the integration service during the sequencing run.
Visible on Record Details screen.
BaseSpace Run ID
Received from the message sent to the integration service.
Hidden
ICA Project ID
Received from the message sent to the integration service.
Hidden
Instrument Run ID
Received from the message sent to the integration service.
Hidden
Run Start Time
Received from the message sent to the integration service.
Hidden
Field Name
Description
Additional Information
Analysis Status
Presets
Started
Completed
Failed
NeedsAttention
Stopped
TimedOut
CompletedWithErrors
Set by the integration service.
Visible on Record Details screen.
Analysis Result Location
Received from the message sent to the integration service.
Visible on Record Details screen.
Library Tube Strip Barcode
Identified by the integration service using information from RunParameters.xml.
Hidden
Field Name | Field Type | Options | Additional Options and Dropdown Items |
PhiX Volume (ul) | Numeric | Read Only | Decimal places displayed: 1 |
1–2% PhiX Spike-In | Toggle Switch | Required Field | Default: No |
Field | Modification | Purpose/Notes |
Final Loading Concentration (pM) |
|
|
Flowcell Type |
|
|
Field | Modification | Purpose/Notes |
Output Folder |
|
|
Field | Modification | Purpose/Notes |
Library Tube Barcode |
|
|
Automation | Setting |
Validate Flowcell Inputs and Validate Analysis Configurations and Register Step Started |
|
Validate Input Count and Validate Analysis Configurations and Register Step Started |
|
Automation | Setting |
Calculate Volumes |
|
The integration includes the following features:
Preconfigured NovaSeq X Series Sequencing v1.1 workflow that maps to lab protocols and instrument runs.
The following steps in the NovaSeq X Series Sequencing v1.1 preconfigured protocol:
Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1)
Make Bulk Pool (NovaSeq X Series Sequencing v1.1)
Dilute and Denature (NovaSeq X Series Sequencing v1.1)
Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1)
AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1)
AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1)
Automatic validation of run setup information before sample sheet generation or planned run creation.
Automated generation of the sample sheet. This sample sheet is used to start the sequencing run on the NovaSeq X Series Control Software through Local run mode.
Automated creation of a planned run on Illumina Connected Analytics (ICA). The NovaSeq X Series Control Software retrieves the planned run for use with the Cloud run mode.
Automated tracking of the NovaSeq X Series sequencing run and parsing of run statistics from BaseSpace Sequence Hub to Clarity LIMS.
Automated tracking of the NovaSeq X Series analysis run and high level analysis summary from BaseSpace Sequence Hub to Clarity LIMS.
Support for configuration of multiple secondary analyses with multiple library prep kits (LPKs) and index adapter kits (IAKs) for the planned run. The analysis is either performed by ICA in Cloud mode or by DRAGEN on-board the instrument in Local mode.
[Optional] Preconfigured Library Prep Validation v2.3.2 workflow used for validation purposes only. The workflow contains a single-step protocol that models the library prep required to produce libraries that are ready for the NovaSeq X Series Sequencing v1.1 workflow. For more information, refer to NovaSeq X Series Integration v1.2.0 User Interaction, Validation and Troubleshooting.
[Optional] Clarity LIMS Product Analytics (CLPA) integration that can be used with ICA. The CLPA integration installation and configuration is required. For more information, refer to Clarity LIMS Product Analytics Integration v1.2.0 Configuration.
Field Name
Field Type
Options
Additional Options and Dropdown Items
Cloud Run ID
Text
Read Only
Hidden
Index 1 Cycles
Numeric Dropdown
Required Field
Custom Entries
Range = 0–20
Presets
0
6
8
Index 2 Cycles
Numeric Dropdown
Required Field
Custom Entries
Range = 0–20
Presets
0
6
8
Read 1 Cycles
Numeric Dropdown
Required Field
Custom Entries
Range = 1–251
Presets
51
101
151
Read 2 Cycles
Numeric Dropdown
Required Field
Custom Entries
Range = 0–251
Presets
51
101
151
Run Mode
Text Dropdown
Required Field
Presets
Local
Cloud
Run Name
Text
Required Field
This section explains how to validate the installation of the Illumina NovaSeq X Series Integration Package v1.2.0.
The validation process involves the following actions:
Running samples through the Library Prep Validation v2.3.2 workflow.
The workflow contains a single-step protocol that models the library prep required to produce libraries tagged with index sequences. At the end of the step, these libraries are routed to NovaSeq X Series Sequencing v1.1 workflow.
Running libraries through the NovaSeq X Series Sequencing v1.1 workflow validates the following items:
Successful sequential step advancement of samples through the steps of the workflow.
Automatic validation of run setup information before sample sheet generation or planned run creation.
Automated sample sheet generation. This sample sheet is used to start the sequencing run on the NovaSeq X Series Control Software through the Local run mode.
Automated creation of a planned run in Illumina Connected Analytics ( ICA). The control software retrieves the planned run. The run is started through the Cloud run mode.
Automated tracking of the NovaSeq X Series sequencing run and parsing of run statistics from BaseSpace Sequence Hub to Clarity LIMS.
Automated tracking of the NovaSeq X Series analysis run and high level analysis summary from BaseSpace Sequence Hub to Clarity LIMS.
The validation steps assume that the following conditions have been met:
The NovaSeq X Series Integration v1.2.0 has a valid BaseSpace Sequence Hub account with an Enterprise or Professional subscription.
NovaSeq X Series Integration Package v1.2.0 is installed and you have imported the default Clarity LIMS configuration.
Analysis configuration templates (ACTs) are created in BaseSpace Sequence Hub (BaseSpace Sequence Hub) for your run. Make sure that the index adapter kit label group is created in Clarity LIMS before selecting it in the ACT. This same label group is used in the Run Library Prep Validation v2.3.2 step. For more information on creating a reagent label group, refer to Add and Configure Labels and Label Groups in the Clarity LIMS (Clarity & LabLink Reference Guide) documentation. For the adapter sequences for your library prep kits, refer to Illumina Adapter Sequences.
You must create the Analysis Configuration Templates (ACTs) that are required for configuring secondary analysis in the NovaSeq X Series Sequencing v1.1 workflows. Create and delete ACTs in BaseSpace Sequence Hub. For instructions, refer to the BaseSpace Sequence Hub Online Help on the Illumina support site).
The following steps set up Clarity LIMS in preparation for running samples through the Library Prep Validation v2.3.2 and NovaSeq X Series Sequencing v1.1 workflows.
On the Configuration tab, under Workflows, activate both the Library Prep Validation v2.3.2 and NovaSeq X Series Sequencing v1.1 workflows.
On the Projects and Samples screen, create a project and add samples to it.
Assign the samples to the Library Prep Validation workflow.
This single-step protocol models the library prep required to produce libraries tagged with index sequences that are ready for the NovaSeq X Series Sequencing v1.1 workflow.
Follow the steps in Library Prep Validation Protocol to run the Library Prep Validation workflow with the following:
Label Group = same as the Index Adapter Kit selected in the ACT that is being used
Sequencing Instrument = NovaSeq X Series
On exit from the step, the Routing Script automation is triggered. This automation assigns samples to the first step of the NovaSeq X Series Sequencing v1.1 workflow, Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1).
The NovaSeq X Series Sequencing v1.1 protocol consists of the following steps:
Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1)
Make Bulk Pool (NovaSeq X Series Sequencing v1.1)
Dilute and Denature (NovaSeq X Series Sequencing v1.1)
Load To Library Tube Strip (NovaSeq X Series Sequencing v1.1)
AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1)
AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1)
Each step contains a script to register the start time (upon step entry) and the time that the step is completed (upon step exit). This information is published to CLPA through ICA. These scripts are used only for CLPA support and can be incorporated as part of an automation that performs other functions.
In Lab View, locate the NovaSeq X Series Sequencing v1.1 protocol. The samples are queued for the Assign Analysis Configuration Template (NovaSeq X Series Sequencing v1.1) step.
Add the samples to the Ice Bucket and select View Ice Bucket.
On the Ice Bucket screen, select Begin Work to start the Validate Sample Names and Retrieve Analysis Configuration Template List and Register Step Started automation.
The automation checks that the sample names do not use restricted characters and are within character limits and retrieves the list of ACTs that you can select. The automation also registers the start time of the step by publishing messages to CLPA through ICA. The part of the script for start time registration is used only for CLPA support.
Select the applicable ACT to assign to the samples. The index adapter kit specified by the ACT must correspond with the label group used in Library Prep Protocol.
[Optional] In Step Details, select Retrieve ACT Information to trigger the Retrieve ACT Information automation.
This automation retrieves ACT information (e.g., Library Prep Kit, Index Adapter Kit, Reference Genome, and so on) and populates the fields in Clarity LIMS. These details are saved to the ACTMetadata.csv file that you can download. If you are not sure of the analysis configuration before starting the sequencing and analysis run, refer to the details in the file.
Select Next Steps to assign the ACT to the samples.
This action triggers the Validate Reagent Labels and Apply Selected ACT to Samples and Set Next Step automation. This automation validates that the indexes applied to the libraries are valid for the selected ACT and assigns samples to it. All samples added to the Ice Bucket (mentioned above) are assigned to this ACT.
Select Finish Step.
In Lab View, locate the NovaSeq X Series Sequencing v1.1 protocol. The samples are queued for the Make Bulk Pool (NovaSeq X Series Sequencing v1.1) step.
Add the samples to the Ice Bucket and select View Ice Bucket.
On the Ice Bucket screen, select Begin Work.
Create a pool of samples as follows.
On the Pooling screen, create a pool by dragging samples into the Pool Creator.
If a control sample was added, it appears under the Sample List and can be pulled into the pool that needs the control.
Enter a name for your pool or accept the default name (Pool #1).
[Optional] If multiple pools are required, select the plus sign (+) next to Pool Creator to create a pool.
[Optional] To remove a pool, select the X in the top right corner of the pool.
Select Record Details to trigger the Validate Analysis Configurations automation.
This automation performs the following checks on the analysis configuration for each pool:
Pooled samples are within the maximum configuration limit.
Pooled samples have the sample type of analysis (Cloud or Local).
Pooled samples that have the same secondary analysis also have the same analysis version and settings.
On the Record Details screen, navigate to the Reagent Lot Tracking section to track the lot information used in the step.
[Optional] Create a new lot. For more information, refer to Add and Configure Reagent Kits and Lots in the Clarity LIMS (Clarity & LabLink Reference Guide) documentation.
In the Step Details area, complete the following required fields:
Number of Lanes to Sequence — Used in volume calculations, to make sure that the volumes are sufficient for the number of times the pool is sequenced. This field is applied to all pools in the step.
Final Loading Concentration (pM) — The final loading concentration of the pool in the flow cell.
Minimum Per Sample Volume (ul) — The minimum volume for each sample. This field is prepopulated with the configured default value (2 µl) and can be edited. If the per sample volume is below the value set in this field, the Calculate Volumes script applies a rounding factor to affected samples so that the volume reaches the minimum volume.
Flowcell Type — The flow cell type that the run uses. This selection affects the computation of volumes in the Calculate Volumes automation that generates the calculation file. Select from the following options:
1.5B
10B
25B
Select Calculate Volumes to trigger the Calculate Volumes automation.
This automation calculates the volumes required for each library to form a pool that has the concentration and volume specified in the Step Details field. It also generates the calculation file in a CSV format and attaches it to the step. Select the file name to download it.
In the Sample Details table, select the pool next to the sample name to view details on the pool composition.
Select Next Steps to trigger the Set Next Step automation.
This automation sets the next step for samples to ADVANCE, which moves them to the Dilute and Denature (NovaSeq X Series Sequencing v1.1) step.
Select Finish Step.
In Lab View, locate the NovaSeq X Series Sequencing v1.1 protocol. The pool of samples queued for the Dilute and Denature (NovaSeq X Series Sequencing v1.1) displays.
Add the pool to the Ice Bucket and select View Ice Bucket.
[Optional] On the Ice Bucket screen, set the number of derivatives to create (placed into the library tube strip) and select Begin Work.
On entry to the step, the Validate Inputs Flowcell Type and Register Step Started automation is triggered. This automation makes sure that the configured flow cell type is valid. The automation also registers the start time of the step by publishing messages to CLPA through ICA. The portion of the script used for start time registration is used only for CLPA support.
If PhiX is used for the run, navigate to the Record Details screen and set 1–2% PhiX Spike-In to Yes.
Select Calculate Volumes to trigger the Calculate Volume automation. This automation sets the following values:
BP Aliquot Volume (ul)
RSB Volume (ul)
NaOH Volume (ul)
TT2 Volume (ul)
PhiX Volume (ul) and Concentration (pM) (if there is a PhiX spike-in)
The automation also generates the calculation file (CSV) and attaches it to the step. This file contains information about the volume of RSB, NaOH, and TT2 to add per working pool. If there is a PhiX spike-in, the file also contains information on the PhiX volume and concentration.
On the Record Details screen, the Reagent Lot Tracking section tracks the NaOH, Resuspension Buffer, and TT2 reagents used in the step. To add and activate reagent lots, refer to Add and Configure Reagent Kits and Lots in the Clarity LIMS (Clarity & LabLink Reference Guide) documentation.
On the Record Details screen, perform the following actions:
In the Reagent Lot Tracking section, select from the active lots displayed in each drop-down list.
In the Sample Details table, make sure that the BP aliquot, RSB, NaOH, and TT2 reagent volume values are populated. The script sets these values and they cannot be edited.
[Optional] In the Sample Details table, select the pool icon to view details of the working pool composition.
In the Files area, select the Calculation File (CSV) to open it and view details on the volumes for each reagent and sample pool used for the dilute and denature process.
Select Next Steps.
Select Finish Step.
In Lab View, locate the NovaSeq X Series Sequencing v1.1 protocol. The pool of samples are queued for the Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) step.
Add the pools to the Ice Bucket and select View Ice Bucket.
On the Ice Bucket screen, select one of the following destination container types:
Library 8-tube Strip
Library 2-tube Strip
Select Begin Work to trigger the Validate Flowcell Inputs and Validate Analysis Configurations and Register Step Started automations.
The Validate Flowcell Inputs automation ensures that the correct container is selected for the flow cell type and that the number of inputs is the same as the number of available wells on the library tube strip. The Validate Analysis Configurations automation is similar to the automation used in the Make Bulk Pool step.
The Register Step Started automation also registers the start time of the step by publishing messages to CLPA through ICA. The portion of the script used for start time registration is used only for CLPA support.
To use multiple ACTs in a single run, repeat Steps 1–4 for each ACT.
The pool of samples for each ACT is then queued for the Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) step to be added to the Ice Bucket.
On the Placement screen, do as follows.
Drag the pools into the Placed Samples area on the right.
Scan or type the barcode of the library tube strip into the container name field.
Select Record Details.
After exiting the Placement screen, the Validate Library Strip Tube Barcode automation makes sure that the library tube strip barcode conforms to the barcode mask. For the Library 8-tube Strip, the barcode mask is LC[0-9]{7}-L[A-Z]{1}1 and an error message displays if the barcode does not match. For the Library 2-tube Strip, the barcode mask is LC[0-9]{7}-L[A-Z]{1}2.
The fields displayed on the Record Details screen are used to create the planned run and generate the sample sheet.
The analysis related information for the planned run is from the ACT associated with the samples and no further analysis configuration is required. Refer to the following table for details.
Fields Displayed on Record Details Screen of Load to Library Tube Strip (NovaSeq X Series Sequencing v1.1) Step
¹ The Local run mode generates the sample sheet. The Run/Analysis configuration is imported manually to the NovaSeq X Series instrument through the sample sheet. The sample sheet is downloaded and imported manually by the user on NovaSeq X Control Software to start the run. The downstream secondary analysis is done using the onboard DRAGEN module.
² The Cloud run mode sends the Run/Analysis configuration to ICA. This configuration is downloaded to the NovaSeq X Series instrument through ICA to start the run. The downstream secondary analysis is done in the cloud.
³ The custom value must correspond to the longest index sequence of the samples in the pools in the library tube strip. Otherwise, the planned run creation fails and an error message displays.
Select Validate Run Setup and Create Planned Run to trigger the automation script. The script performs the following actions:
Validates the parameters entered on the Record Details screen.
Creates the planned run and sends it to ICA when the Run Mode is Cloud.
Generates the sample sheet and attaches it to the placeholder in the Files area on the Record Details screen for all Run Mode types.
Select Next Steps.
On the Assign Next Steps screen, the next step for the pooled samples is set to the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step.
Select Finish Step to advance the pooled samples to the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step. For more information on how to start the sequencing run for different run modes, refer to NovaSeq X Series Integration v1.2.0 Configuration.
This protocol contains the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step. This step is fully automated.
The integration starts the step automatically and data from the run is parsed back into Clarity LIMS. User interaction is not required, but you can review the stages of the step in Clarity LIMS.
Read summary metrics are recorded for the library pool. After the run is complete, open the step and review these metrics in the Step Details section and the Sample Details table.
This protocol contains the AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) step. This step is fully automated.
The integration starts the step automatically and data from the run is parsed back into Clarity LIMS. User interaction is not required, but you can review the stages of the step in Clarity LIMS. If the run analysis is successful, the integration completes the step automatically.
After the analysis run is complete, open the step and review the following values in the Step Details section:
Analysis Status
Analysis Result Location
Log
After the analysis is complete, download the demultiplexing results and analysis results summary files from the file placeholders. Retrieve detailed analysis results from ICA.
The demultiplexing results file contains the following demultiplexing statistics for each sample:
Number of reads
Number of perfect index reads
Number of mismatch index reads
The analysis result summary file provides a summary of what analysis workflows were performed. This file also identifies the total number of samples analyzed and how many were completed or failed for each analysis.
At this point, the entire NovaSeq X Series Integration workflow is fully validated.
If an automation trigger does not appear to run its corresponding scripts, refer to the Troubleshooting Automation section in the Clarity LIMS (API & Database) documentation.
If the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step does not start, Clarity LIMS is likely not receiving sequencing run events from BaseSpace Sequence Hub correctly. Check for the following issues:
Make sure that the NovaSeq X Control Software is configured properly as follows.
Before you start the run, in the NovaSeq X Control Software, select the appropriate BaseSpace Sequence Hub region in the Hosting location drop-down list.
Under Run Settings, check the boxes next to Cloud run monitoring and Cloud run storage.
If there are issues after using the configuration settings on the NovaSeq X Control Software, contact Illumina Support.
If the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step starts, but does not complete, do as follows.
Log in to the default user account and use one of the following methods to open the in progress step in Clarity LIMS:
In Lab View, find the step in the Recent Activities pane.
Search for the step in Clarity LIMS using the library tube strip barcode as the search term.
On the Record Details screen, the Sequencing Log multiline text field contains logging information.
If unable to reach the Record Details screen, or if the Sequencing Log field does not contain enough information to resolve the issue, contact Illumina Support. Provide the relevant information from the troubleshooting steps already performed.
If you receive an error when creating a planned run, check the log message in the Load to Library Tube Strip step to identify the issue. If you cannot correct the issue, contact Illumina Support.
Incompatible Analyses in a Planned Run
Only compatible analysis versions should be combined in a single planned run. When incompatible analysis versions are combined, an error log message displays. An example of the error log message is shown below.
To resolve this error, check the ACTs that were used for the run and only select the ACTs that are compatible with the planned run.
The Validate Analysis Configuration automation check is in the Make Bulk Pool (upon pooling) and Load to Library Tube Strip (upon step entry) steps. If a failure happens, an error message displays and the step can be aborted, or you might be prevented from continuing the step. For NovaSeq X Series Integration v1.2, the configuration limits for the Cloud and Local modes are as follows.
Cloud — Maximum of seven applications and genome configurations (and, if needed, the BCL Convert Only application). Cloud mode also allows up to eight different library prep kits for each application and genome configuration.
Local — Maximum of three applications and genome configurations (and, if needed, the BCL Convert Only application). Local mode also allows up to eight different library prep kits for each application and genome configuration.
This check can fail for the following reasons:
The analysis configuration after library pooling or during the planned run creation exceeds the configuration limit.
Resolve this error as follows.
If the error occurs on the Make Bulk Pool step, reduce the number of analysis configurations for a pool. To reduce the number, reorganize the samples for the pooling step (eg, by pooling samples that have similar configurations together).
If the error occurs on the Load to Library Tube step, reduce the number of analysis configurations for the planned run by reorganizing the samples for multiple runs instead of a single run.
ACTs of samples in the same pool or planned run must have the same run mode (Local or Cloud).
Resolve this error as follows.
Abort the step and remove samples with ACTs that have conflicting run modes from the Ice Bucket.
Make sure that all remaining samples in the Ice Bucket have ACTs with the same run modes.
Select Begin Work to continue the step.
ACTs of samples in the same pool or planned run that have the same secondary analysis must have the same analysis version.
Resolve this error as follows.
If the error occurs on the Make Bulk Pool step, pool the samples again. Make sure that samples with the same secondary analysis, but conflicting analysis versions, are in different pools.
If the error occurs on the Load to Library Tube step, reorganize the samples for the planned run. Make sure that samples with the same secondary analysis, but conflicting analysis versions, are in different runs.
ACTs of samples in the same pool or planned run that have the same secondary analysis must have the same analysis settings. The analysis setting fields can differ for a different secondary analysis and are configured in BaseSpace Sequence Hub when the ACT is created.
Resolve this error as follows.
If the error occurs on the Make Bulk Pool step, pool the samples again. Make sure that samples with the same secondary analysis, but conflicting analysis settings, are in different pools.
If the error occurs on the Load to Library Tube step, reorganize the samples for the planned run. Make sure that samples with the same secondary analysis, but conflicting analysis settings, are in different runs.
Last Updated: November 2024
Release Date: February 2024
Document Version: 2
These release notes describe the key changes to software components for the Clarity LIMS NovaSeq X Series Integration Package v1.2.0.
Refer to Compatibility under Instruments & Integrations.
Supports analysis tracking at the metaworkflow level for analysis status and high level analysis summary results.
Supports the new 1.5B and 25B flow cell types. The B3 flow cell has been renamed to the 10B flow cell.
Uses the SecureString type for improving the security of values stored in the AWS param store.
Includes the following improvements to the NovaSeq X Series Sequencing v1.1 protocol:
Removes the unused Output Folder step custom field from the Load to Library Tube Strip step.
Renames the Library Tube barcode step custom field to Library Tube Strip Barcode in the AUTOMATED - Sequencing Run step.
None
The analysis step tracks only the first run, but not subsequent queued runs.
The Local run mode does not support custom library prep and index adapter kits.
The Assign Analysis Configuration Template step expects the input samples to be unpooled libraries.
On the Make Bulk Pool step, the log displays a warning when the Calculate Volume automation is triggered and at least one pool consists of multiple inputs. This issue is caused by the output custom field being reset multiple times at the end of the automation. This issue does not affect the Calculate Volume automation functionality.
Sample sheet and planned run generation will fail if any of the samples in the pools has been assigned QC flag in prior steps before entering the Load to Library Tube Strip step.
Used for creating a planned run in ICA.
Sample and library names must use only alphanumeric, dash, or underscore characters. Invalid characters can cause a sample sheet validation failure in the Load to Library Tube Strip step.
If PhiX control sample is required for the run, the configuration setting shall be performed in the Step 3: Dilute and Denature with the 1–2% PhiX Spike-In toggle switch.
Do not add samples to the Ice Bucket or start or complete the AUTOMATED - Sequencing Run (NovaSeq X Series Sequencing v1.1) step. The integration does this automatically.
Do not add samples to the Ice Bucket or start or complete the AUTOMATED - Analysis Run (NovaSeq X Series Sequencing v1.1) step. The integration does this automatically.
Version
Changes
2
Updated Compatibility section to reference Compatibility matrix table.
Updated Known Limitations section.
1
Initial release.
Field
Description
Run Name
Enter the experiment name. Only alphanumeric characters, dashes, and underscores are permitted. No spaces.
Run Mode
Presets
Local¹
Cloud²
Read 1 Cycles
Presets
151
101
51
Type a custom value
Read 2 Cycles
Presets
151
101
51
Type a custom value
Index 1 Cycles
Presets
0
6
8
Type a custom value³
Index 2 Cycles
Presets
0
6
8
Type a custom value³
The selected Run Mode must correspond to the ACT type.
