> For the complete documentation index, see [llms.txt](https://help.connected.illumina.com/llms.txt). Markdown versions of documentation pages are available by appending `.md` to page URLs; this page is available as [Markdown](https://help.connected.illumina.com/dragen-stratamap/dragen-spatial-transcriptome/analysis-inputs/sample-sheet.md).

# Sample Sheet

A sample sheet is required for each analysis with the DRAGEN Spatial Transcriptome analysis software. A sample sheet is a comma-separated value (\*.csv) file format used by Illumina instruments, platforms, and analysis pipelines to store settings and data for sequencing and analysis. The DRAGEN Spatial Transcriptome analysis software is compatible with sample sheet v2. For general information on sample sheet v2, refer to [Illumina Connected Software - Sample Sheet](https://help.connected.illumina.com/run-set-up/overview).

{% hint style="info" %}
The analysis fails if the sample sheet requirements are not met. We recommend using the BSSH Run Planning tool to generate the sample sheet (see [Run Planning](/dragen-stratamap/dragen-spatial-transcriptome/run-setup/run-planning.md)).
{% endhint %}

### Sample Sheet Format

The following sample sheet requirements describe required and optional fields for DRAGEN Spatial Transcriptome analysis.

Use the following steps to generate a valid sample sheet:

1. Use the BaseSpace Sequencing Hub Run Planning tool to generate an example sample sheet ([Run Planning](/dragen-stratamap/dragen-spatial-transcriptome/run-setup/run-planning.md)) or download the sample sheet v2 template for DRAGEN Spatial Transcriptome analysis.
2. If using a template, within the Sequencing Settings section, enter the following required parameters:

## \[Sequencing\_Settings] Section

<table><thead><tr><th>Sample Parameter</th><th width="215">Required</th><th>Details</th></tr></thead><tbody><tr><td>LibraryPrepKits</td><td>Required</td><td>Accepted value: IlluminaSpatialLibraryPrep</td></tr></tbody></table>

\
3\. In the BCL Convert Settings section, enter the following required parameters:

## \[BCLConvert\_Settings] Section <a href="#bclconvert_settings-section" id="bclconvert_settings-section"></a>

| Sample Parameter | Required | Details               |
| ---------------- | -------- | --------------------- |
| SoftwareVersion  | Required | Accepted value: 4.4.0 |

4. In the BCL Convert Data section, enter the following parameters for each sample.

## \[BCLConvert\_Data] Section

| Sample Parameter | Required                                                         | Details                                                                                                                                                                                  |
| ---------------- | ---------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- |
| Lane             | Only for NovaSeq 6000 XP, NovaSeq 6000Dx, or NovaSeq X workflows | Indicates which lane corresponds to a given sample. Enter a single numeric value per row. Cannot be empty; the analysis fails if the Lane column is present without a value in each row. |
| Sample\_ID       | Required                                                         | Must match a Sample\_ID listed in the Spatial Data section.                                                                                                                              |
| Index            | Required                                                         | Set to "na" for each sample.                                                                                                                                                             |

5. In the Cloud Spatial Settings section, enter the following parameters:

## \[Cloud\_Spatial\_Settings] Section

<table><thead><tr><th width="327">Setting Parameter</th><th>Required</th><th>Details</th></tr></thead><tbody><tr><td>SoftwareVersion</td><td>Required</td><td>The DRAGEN Spatial Transcriptome software version.</td></tr><tr><td>StageAllSettings</td><td>Required</td><td>Set to TRUE for DRAGEN Spatial Transcriptome.</td></tr><tr><td>GridBinSize</td><td>Required</td><td>Array of integers delimited by colons, e.g., 5:10:25 for bin sizes 5, 10, and 25.</td></tr><tr><td>CellBoundaryExpansionDistance</td><td>Required</td><td>Set the cell expansion distance from the cell nucleus, in μm.</td></tr><tr><td>MapAlignOutFormat</td><td>Required</td><td>Recommendation is to set this to "None". Set to "bam" to generate BAM files for each sample. BAM file generation will incur additional storage and compute costs.</td></tr><tr><td>Genome</td><td>Required</td><td>Provide the URN to the reference genome.</td></tr></tbody></table>

6. In the Cloud Spatial Data section, enter the following parameters:

## \[Cloud\_Spatial\_Data] Section

| Sample Parameter | Required | Details                                                                               |
| ---------------- | -------- | ------------------------------------------------------------------------------------- |
| Sample\_ID       | Required | Must match the Sample\_ID listed in the BCL Convert Section.                          |
| ImageFile        | Required | The file name of the registered image file generated by Spatial Image Aligner.        |
| ImageLocation    | Required | The URN of the registered image file generated by Spatial Image Aligner in ICA.       |
| Substrate ID     | Required | ID of the substrate used for the spatial experiment. Only one substrate is supported. |

7. In the Cloud Settings section, enter the following parameters:

## \[Cloud\_Settings] Section

| Setting Parameter        | Required     | Details                                                                                              |
| ------------------------ | ------------ | ---------------------------------------------------------------------------------------------------- |
| GeneratedVersion         | Not Required | The cloud GSS version used to create the sample sheet. Optional if manually updating a sample sheet. |
| CloudWorkflow            | Not Required | ica\_workflow\_1                                                                                     |
| Cloud\_Spatial\_Pipeline | Required     | This value is a universal record number (URN).                                                       |

8. In the Cloud Data section, enter the following parameters:

## \[Cloud\_Data] Section

| Parameter           | Required     | Details                                                      |
| ------------------- | ------------ | ------------------------------------------------------------ |
| Sample\_ID          | Not Required | The same sample ID used in the Cloud\_Spatial\_Data section. |
| ProjectName         | Not Required | The BaseSpace project name.                                  |
| LibraryName         | Not Required | For spatial, this is the same as the sample ID.              |
| LibraryPrepKitName  | Required     | IlluminaSpatialLibraryPrep                                   |
| IndexAdapterKitName | Not Required | IlluminaSpatial                                              |


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