Sample sheets can be edited and FASTQ Generation analysis requeued for iSeq, MiniSeq (updated to Control Software 2 and Windows 10), MiSeq, HiSeq, NextSeq 500/550 (updated to Control Software 4 and Windows 10) and NovaSeq runs uploaded to BaseSpace Sequence Hub. The sample sheet is a comma-separated values (*.csv) file that stores information required to set up, perform, and generate demultiplexed FASTQs from a sequencing run. Illumina recommends using the Windows-based Illumina Experiment Manager software to create the sample sheets, as it guides users through the process and will check for errors.
There are various reasons to requeue a FASTQ Generation analysis. For example, if indexes were entered incorrectly into the sample sheet (e.g., reverse complement), the indexes must be corrected and the data demultiplexed again. Techniques for troubleshooting demultiplexing issues can help users diagnose demultiplexing issues. Requeuing can also be used to turn adapter trimming on or off, or to reformat the sample sheet to remove any illegal characters. Examples of illegal characters are spaces or any characters other than alphanumerics, dashes, or underscores in the Sample ID field of the sample sheet.
- MiniSeq and NextSeq 500/550 runs set up using the BaseSpace Sequence Hub Preptab require different steps to edit run setup and requeue an analysis.
- Only the owner of the run can requeue an analysis. It is possible to transfer ownership of a run, if necessary. A shared run does not provide the necessary permissions needed to requeue analysis.
- There is a limit of five requeue attempts for users of BaseSpace Sequence Hub. If assistance with additional requeues is needed, email Illumina Technical Support, and be prepared to share your run ID and sample sheet. To find the run ID, open the run in BaseSpace, and look in the web browser address bar for the numerical value after: https://basespace.illumina/com/run/
- After a sample sheet is edited, only the originally uploaded file shows up in the Sample Sheet tab for the run, but any changes made are used for analysis.
Use BaseSpace Sequence Hub to edit a sample sheet and requeue:
- After logging in to your BaseSpace account, select the RUNS tab at the top of the page.
- Select the blue hyperlink of the run you wish to requeue.
- Select the hourglass symbol drop-down menu, then select Requeue and then Sample Sheet.
Note: this option will be grayed out if you are not the owner of the run.
- Select Load Original to load the original sample sheet for editing. Any errors detected in the sample sheet will be displayed above the editing window. Edits can be directly entered or copy/pasted from a plain text editor, such as Notepad, to maintain comma-separated values. Edits cannot be copied from a spreadsheet program, like Excel. When edits are complete, select the blue Queue Analysis button. If the Queue Analysis button is not active, check for and resolve any validation errors.
- The status on the run summary page will display “Analyzing” when the analysis is ongoing and will change to “Complete” when the requeued analysis is complete.