# Sample Sheet Troubleshooting ## General troubleshooting for a failed sample sheet: In DRAGEN TruSight Oncology 500 Analysis Software, the analysis fails if a sample sheet is invalid. If an invalid sample sheet in suspected, log files can help troubleshoot a failed analysis. Use the following steps to find the log file for the sample sheet: 1. Navigate to the following location `//Logs_Intermediates/SamplesheetValidation.` 2. Open the `SamplesheetValidation-.log` file 3. Find a line with the following: `SampleSheetValidationTask:NA:1 exited with return code 1 which has not been declared as a valid return code.` 4. Search for errors in the sample sheet validation log and compare with the guidelines and warnings in [Sample Sheet Requirements](/tso500/dragen-tso-500-guides/dragen-tso-500-v2.6/run-planning/sample-sheet-requirements.md) and the following tables. | Failure Type | Action | | ---------------------------------------------------- | -------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | | Sample Sheet not found | Verify that `SampleSheet.csv` is present at the top level of the run folder with the name "SampleSheet.csv". If the sample sheet is in a different location, supply the sample sheet using the `--sampleSheet` option | | Indexes are not valid for the sequencer and/or assay | See Valid indexes for assay and instrument combinations for correct indexes for the sequencer and assay. | | Pair\_ID is not unique | Pair\_ID column is required in the TSO500S\_Data section of the sample sheet, which pairs at most one RNA and one DNA sample together for analysis. If the sample does not have a pair, use a unique pair ID for single samples. | | Sample Sheet is not in v2 format | Verify that the format of the sample sheet is v2. v1 sample sheet is not compatible with DRAGEN TruSight Oncology 500 Analysis Software. | | Analysis does not run | Verify the analysis starts from the run folder, and BCLs or FASTQs are in the correct locations as outlined in Starting From BCL Files and Starting From FASTQ Files respectively. | ## Indexes included in Library Prep Kit: | Assay | Index Set ID | | ---------- | ------------------------------------------------------------ | | TSO 500 | | | TSO 500 HT | | | TSO 500 v2 | | ## Troubleshooting BCL issues | Failure Type | Action | | -------------------------------------- | -------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | | Lane Column without Values | Ensure that the column is completed. If lane is not applicable to the run, delete the column. | | Format of v2 sample sheet is incorrect |

Verify that the following sections and fields are present in the sample sheet and follow the individual rules in Sample Sheet Requirements

\[BCLConvert\_Settings]
- SoftwareVersion
- AdapterRead1
- AdapterRead2
- AdapterBehavior
- MinimumTrimmedReadLength
- MaskShortReads

\[BCLConvert\_Data]
- Sample\_ID
- index
- index2

\[TS0500S\_Data]
- Sample\_ID
- Index\_ID
- Sample\_Type
- Pair\_ID
- Sample Feature (Optional)

| | HRD analysis is missing | Verify that HRD is in the Sample Feature column in the sample sheet. Refer to [Sample Sheet Requirements](/tso500/dragen-tso-500-guides/dragen-tso-500-v2.6/run-planning/sample-sheet-requirements.md) for more information. | ## Troubleshooting FASTQs issues | Failure Type | Action | | --------------------------------------------- | ------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | | Sample\_ID and/or Sample\_Type is not present | Verify that the sample sheet has columns and values for Sample\_ID and Sample\_Type. | | Unique sample IDs | Verify that the Sample\_IDs are unique in the sample sheet. | | Format of v2 Sample Sheet is incorrect |

Verify that the following sections and fields are present in the sample sheet and follow the individual rules in Sample Sheet Requirements.
\[TS0500S\_Data]
- Sample\_ID
- Index\_ID
- Sample\_Type
- Pair\_ID

- Sample Feature (Optional)
Verify when FASTQs were generated using the HRD add-kit (Not available in Japan), Sample Feature is added to those DNA Samples. Refer to Sample Sheet Requirements for more information.

| | Incorrect folder structure | Verify that the FASTQ files are in the correct structure. Refer to [Starting From FASTQ Files](/tso500/dragen-tso-500-guides/dragen-tso-500-v2.6/launching-analysis/run-dragen-tso-software/command-line-options.md#starting-from-fastq-files) for more information. | | Invalid FASTQ input files | If the FASTQs are invalid, start TSO 500 analysis from BCL files. | | HRD analysis missing | Make sure that HRD is in the correct column in the sample sheet. | --- # Agent Instructions: Querying This Documentation If you need additional information that is not directly available in this page, you can query the documentation dynamically by asking a question. Perform an HTTP GET request on the current page URL with the `ask` query parameter: ``` GET https://help.connected.illumina.com/tso500/dragen-tso-500-guides/dragen-tso-500-v2.6/troubleshooting/sample-sheet-troubleshooting.md?ask= ``` The question should be specific, self-contained, and written in natural language. The response will contain a direct answer to the question and relevant excerpts and sources from the documentation. Use this mechanism when the answer is not explicitly present in the current page, you need clarification or additional context, or you want to retrieve related documentation sections.