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    • References
      • Configure Multiple Identical netPathPrefixSearch Values
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On this page
  • Overview
  • MiSeqDx Assays Support
  • Configured Master Step Fields/Step UDFs
  • Submitted Sample Global Fields/UDFs
  • Script Parameters and Usage
  • Support for Container Types
  • Sample Sheet Data
  • Header Section
  • Manifests Section
  • Reads Section
  • Settings Section
  • Data Section
  • File Format and Contents
  • Configuration Options
  • Rules and Constraints

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  1. Instruments & Integrations
  2. MiSeqDx

MiSeqDx Sample Sheet Generation (v1.11.0 and later)

Package Version: BaseSpace Clarity LIMS MiSeqDx (v1.11.0 and later)

Overview

The Illumina MiSeqDx Integration Package allows for automatic generation of a sample sheet to be used with the MiSeqDx instrument software. The format of this sample sheet is designed for the instrument when it is running in Diagnostic mode.

Note

  • If you are running the MiSeqDx instrument in Research Use Only (RUO) mode, see Sample Sheet Generation section in the Configuration guide for MiSeq version-of-interest.

  • MiSeqDx does not support bcl2fastqv2 sample sheet generation.

How sample sheet generation works

Sample sheet generation is configured on the step prior to the sequencing run – Denature, Dilute and Load Sample, which is the step where samples are placed on the flow cells or reagent cartridges that will be placed in the instrument.

The sample sheet is generated by means of a script, which the lab user initiates by clicking a button on the Record Details screen of the step. This generates a sample sheet file for the container loaded during the step, where the name of the sample sheet will be

<reagent cartridge barcode ID>.csv

The user then downloads the sample sheet from the LIMS and uploads it to the instrument software.

MiSeqDx Assays Support

The following assays are supported by the MiSeqDx sample sheet generation script:

  • CF 139-Variant Assay

  • CF Clinical Sequencing Assay

Configured Master Step Fields/Step UDFs

Sample sheet format is controlled via master step field / step user defined field (UDF) configuration, where key step fields are pre-populated with values specific to each protocol step. These values are not editable, and their configuration should not be modified.

The fields listed in the following table are available on the Denature, Dilute and Load Sample step and will be placed into the sample sheet.

Field Name

Field Type

Required?

Notes

Experiment Name

Text

No

Entered by the user

Workflow

Text

Yes

Value set to Amplicon for CF 139-Variant Assay and CF Clinical Sequencing Assay

Description

Text

No

Entered by the user

Assay

Text

No

Configured with the following preset values

  • CF 139-Variant

  • CF Clinical Sequencing

  • /ul>

Amplicon Workflow Type

Text

No

Configured with the following preset values

  • CF139VARIANTASSAY

  • CFCLINICALSEQUENCINGASSAY

Application

Text

Yes

Configured with the following preset values

  • CF 139-Variant Assay

  • CF Clinical Sequencing Assay

PhiX Control added?

Check box

No

Default set to false

VariantCaller

Text

No

Value set to Starling, a legacy variant caller

Variant Min Quality Cutoff

Numeric

No

Value set to 100

GenomeFolder

Text

Yes

Required for secondary analysis

Control CAT Manifest

Text

No

Preset value CFTRManifest.txt

Custom CAT Manifest

Text

No

Manifest file for custom assay

Read 1 Cycles

Numeric

Yes

Configured with range 0-1000. Value set to 151

Read 2 Cycles

Numeric

Yes

Configured with range 0-1000. Value set to 151

Submitted Sample Global Fields/UDFs

Field Name

Field Type

Required?

Notes

Reference Genome

Text

No

Optionally used to populate the GenomeFolder value for individual samples in the sample sheet.

Control?

Text

No

Used to indicate a control sample that is represented as a submitted sample in the LIMS.

Script Parameters and Usage

Parameter

Description

Required?

Notes

u, username

LIMS username

Yes

p, password

LIMS password

Yes

i, processURI

LIMS process URI

Yes

c, csvFileLimsIds

Sample sheet CSV file LIMS ID

Yes

May be provided multiple times

e, errorLogFileName

Log file name

Yes

l, useProjectLimsID

Project LIMS ID will be used instead of project name in the Project column of the sample sheet

No

Accepted values: true or false. Provide with quotes e.g. -l 'true'

s, useSampleLimsID

Should be set to true Sample LIMS ID will be used instead of sample name in the SampleName column of the sample sheet

No

a, appendLimsID

Should be set to false LIMS ID of the protocol step will be appended to sample names in the SampleName column of the sample sheet.

No

Accepted values: true or false. Provide with quotes e.g. -a 'true'

Usage

Below is the default command line that ships with the Denature, Dilute and Load Sample (CF 139-Variant Assay) step.

bash -c "/opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -u {username} -p {password} -i {processURI:v2} script:setUDF -f 'Progress' -t '//input/@uri->//sample/@uri' -v 'Library ready for sequencing' && /opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/miseqdx/miseqdx-extensions.jar -u {username} -p {password} -i {processURI:v2} script:generate_miseqdx_sample_sheet -c {compoundOutputFileLuid1} -e {compoundOutputFileLuid2} -useSampleLimsID true && /opt/gls/clarity/bin/java -jar /opt/gls/clarity/extensions/ngs-common/v5/EPP/ngs-extensions.jar -u {username} -p {password} -i {processURI:v2} script:labelNonLabeledOutputs -l 'NoIndex'"

Support for Container Types

Single well container types, and all one-dimensional container types with both numeric rows and numeric columns, are supported.

Sample Sheet Data

The following table lists and describes the fields included in the MiSeqDx sample sheet.

Note the following:

  • If no upstream pooling is detected, LIMS will populate the sample sheet with the SampleID and SampleName of the submitted sample. Other fields are populated with data from the samples that were input to the step (i.e. derived samples).

  • If upstream pooling is detected, LIMS will populate the sample sheet with the first upstream pooled inputs found – not with the submitted sample or step input fields.

Control samples may be one of the following:

  • Built-in BaseSpace Clarity LIMS control samples

  • Submitted samples with field / UDF Control? set to true

Header Section

Field Name

Description

Required?

Notes

WMFileVersion

Illumina Worklist Manager Version number.

No

Date

The date the sample sheet was generated.

No

Workflow

Master step field/Step UDF of the same name.

Yes

Application

Populated with CF 139-Variant Assay or CF Clinical Sequencing Assay.

Yes

Assay

Master step field/Step UDF of the same name.

No

Description

Master step field/Step UDF of the same name.

No

Chemistry

The recipe fragments used to build the run-specific recipe.

Yes

Populated with Amplicon

Manifests Section

If a value is provided for only a single CAT manifest file, then all samples in the sample sheet will be given the designation (A) associated with that CAT type.

Field Name

Description

Required?

Notes

A

Master step field/Step UDF Control CAT Manifest. Typically a path to a file.

No

Must be a real path. Convention indicates this is the Control CAT. Control samples will be given the designation A in the Data section.

B

Master step field/Step UDF Control CAT Manifest. Typically a path to a file.

No

Must be a real path. Convention indicates this is the Control CAT. Control samples will be given the designation B in the Data section.

Reads Section

Field Name

Description

Required?

Notes

ℹ Read cycle entry is listed beneath the [Reads] heading, in the first column of the spreadsheet.

Master step field/Step UDF Read 1 Cycles

Yes

Index reads are determined by the MOS, based on the indexes on the inputs.

ℹ Read cycle entry is listed beneath the [Reads] heading, in the first column of the spreadsheet.

Master step field/Step UDF Read 2 Cycles

No

Index reads are determined by the MOS, based on the indexes on the inputs.

For single read, one entry is listed beneath the [Reads] heading, in the first column of the spreadsheet. For paired end, two entries are listed.

Settings Section

Field Name

Description

Required?

Notes

AmpliconWorkflowType

Master step field/Step UDF Amplicon Workflow Type.

No

Populated with values CF139VARIANTASSAY or CFCLINICALSEQUENCINGASSAY

VariantCaller

Master step field/Step UDF of the same name.

No

VariantMinimumQualCutoff

Master step field/Step UDF Variant Min Quality Cutoff.

Yes

Data Section

Field Name

Description

Required?

Notes

Sample_ID

Populated with the LIMS ID of the sample if pooled, or the LIMS ID of the submitted sample if not pooled.

Yes

Sample_Name

Populated with the sample name if pooled, or the submitted sample name if not pooled.

Always present

Sample_Plate

Name of the Container that the Sample resides in as recorded in the LIMS.

Always present

Sample_Well

Container well location of the sample. If a sample is part of a pool, this will list the well location of the sample that was added to the pool.

Always present

Sample_Project

The name of the project in the LIMS, that the sample belongs to.

Always present

Control

Blank for a normal sample. Populated with value positive when Positive Control for MiSeqDx has been added to the pool. Populated with value negative when Negative Control for MiSeqDx has been added to the pool.

No

Controls must have an index. Controls in these cases look just like the other samples.

index

Determined from the reagent label. Uses the Sequence attribute value from Index Reagents. Dual index reagents will contain a hyphen-separated DNA sequence; this field will use the first half of that value.

Yes, if more than 1 input

I7_Index_ID

Determined from the name of the index reagent type. Dual index names will be hyphen-separated; this field will use the first half of that value.

Yes, if more than one input.

index2

Determined from the reagent label. Uses the Sequence attribute value from Index Reagents. Dual index reagents will contain a hyphen-separated DNA sequence; this field will use the second half of that value.

No

I5_Index_ID

Determined from the name of the index reagent type. Dual index names will be hyphen-separated; this field will use the second half of that value.

No

Manifest

A

Yes

GenomeFolder

If master step field/step UDF Use submitted sample details for Genome Folder location is true, this is populated with the value of submitted sample global field/UDF Reference Genome. Otherwise, populated with the value of master step/step UDF GenomeFolder.

Yes, if Use submitted sample details for Genome Folder location is true

Folder path for ReferenceGenomes used for secondary analysis.

Description

No

File Format and Contents

This section outlines the format and contents of the generated sample sheet and associated log file.

When validating the installation of your integration, refer to this information to ensure that the sample sheet and log files are correctly generated.

MiSeqDx sample sheet

  • The file is a comma-separated file.

  • The file contains the following sections:

    • Header

    • Manifests

    • Reads

    • Settings

    • Data

  • The file is populated with data from the samples in the step. If pooled, each sample in the pool is represented as a separate, demultiplexed entry.

  • The entries are sorted by SampleWell and by SampleID.

  • The data section of the file contains 11 columns.

MiSeqDx sample sheet log file

  • The file is in HTML format.

  • The file contains logging information and a success message if sample sheet generated successfully.

Configuration Options

Enabling unique FASTQ file names

To enable unique FASTQ file names per sequencing run, the EPP command on the process type must be configured to use the following parameter options:

  • -useSampleLimsID – ensures unique entries in the SampleName column by using the sample LIMS ID instead of its name

  • -appendLimsID – ensures unique names per run by appending the LIMS ID of the current step

Rules and Constraints

  • The step on which this script runs must be the step in which samples are placed on the flow cell(s) or reagent cartridge(s).

  • The contents of the sample sheet are ordered by SampleWell and then ordered by SampleID.

  • Project and sample names in the sample sheet cannot contain illegal characters. Characters not allowed are the space character and the following: ? ( ) [ ] / \ = + < > : ; " ' , * ^ | &

  • Illegal characters will be replaced with an underscore "_"

  • The destination container type (flow cell or reagent cartridge) must be must be either single well or a one-dimensional container type with both numeric rows and numeric columns. Back to top

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For details, see and sections below.

Accepted values: true or false. Provide with quotes e.g. -s 'true' See Enabling unique FASTQ file names in

If script parameter useSampleLimsID is provided on the command line, the LIMS ID of the sample will be used instead of the name. The additional -a command line option appends the LIMS ID to the end of this value, e.g. Sample1-1234 See .

Value determined by the entries in the .

For more information, see .

Sample Sheet Data
File Format and Contents
Script Parameters and Usage
Configuration Options
Script Parameters and Usage
Manifests Section