How to Create a Sample Sheet in Local Run Manager v2 and v3

For MiSeq Control Software (MCS) v3.1 and later, sample sheet creation should be performed through Local Run Manager (LRM).

Considerations:

• Illumina recommends creating the sample sheet prior to preparing sample libraries.

• Before starting the run, make sure the sample sheet is accessible to the instrument.

  • Copy the sample sheet to a network location accessible to the instrument or transfer via a USB drive onto the instrument. - If copying from network or USB drive, it is best practice to store the file locally and reference the local file location during run setup.

• When the run begins, the software copies the sample sheet from the designated sample sheet folder to the root of the MiSeq Output folder. At the end of the run, the sample sheet is used for secondary analysis performed by LRM.

Follow the instructions below to create a sample sheet:

1. Connect to LRM via a Chromium web browser using the domain https://localhost/#/home

2. Select Create Run

3. Select the desired workflow (For example: Generate FASTQ)

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1. Enter the desired Run Name

2. Select the desired Library Prep Kit from the drop down menu

• If using a non-Illumina library prep kit or a kit type that is not listed, select 'Custom'

1. Select the desired Index Kit from the drop down menu

• If using a non-Illumina library prep kit or a kit type that is not listed, select 'Custom'

1. Select the number of Index Reads (0, 1, or 2)

2. Select the Read Type (Single Read or Paired End)

3. Enter the desired Read Lengths for Read 1 and Read 2

• Note: The Read Lengths for the Index reads will auto-populate based on the number of bases entered into the index columns in the sample sheet.

• If selecting a specific kit from the drop down menu, the index lengths will be auto-populated based on predetermined settings of the kit.

1. Enter the desired number of rows needed and select the '+' symbol to add the rows for the appropriate number of samples

• In the example screenshot below, 17 rows are added for a total of 18 rows.

1. Enter the sample names into the Sample ID column by either copying and pasting from an excel file or entering each sample name individually

• If also using BaseSpace, Illumina recommends using unique sample IDs to avoid potential data aggregation during downstream analyses

1. Enter the appropriate indexes

• If using a specific kit from the drop down menu, the indexes can be selected from the drop down menu.

• Alternatively, if the indexes (or well numbers) are stored in an excel file, that column can be copied and pasted into LRM OR selected by hand.

  • For example, if a previously created sample sheet lists out well A01-H12, the well IDs can be copied and pasted directly into LRM and the associated indexes will populate accordingly.

1. Select Export Sample Sheet and/or Save Run

The sections listed below are "optional settings" that may be desired but are not required by the software to complete run setup:

1. Run Description

• Enter a run description if desired

1. Sample Description

• Enter a sample description if desired

1. Sample Project

• Enter the desired project name here for samples to be associated with said project once demultiplexing completes on BaseSpace

1. Module-Specific/Advanced Settings

• For example, adapter trimming settings can be entered here if a custom kit is used.

  • ​​​​​​​Note: If using a kit from the drop-down menu, the adapter trimming settings for that kit will be applied automatically and do not need to be entered by the user

For additional information, see the Local Run Manager v3 Software Guide and contact Illumina Technical Support if additional assistance is needed.

For any feedback or questions regarding this article (Illumina Knowledge Article #8106), contact Illumina Technical Support [e-mail].