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How to demultiplex a sequencing run without using either the i7 or the i5 index using BCL Convert

If a run is set up with single-indexed and dual-indexed samples in the same run, it is possible to demultiplex the single-indexed samples using BCL Convert, use the following steps. Or more generally, to demultiplex without using either the i7 (index 1) or i5 (index 2), use the following steps. This process is also referred to as index masking (the index that is not use is masked).
To demultiplex the run without i5 index The Sample sheet must have the following options:
  • OverrideCycles with the N# for the i5. For example: OverrideCycles,Y151;I8;N8;Y151
  • The Index2 column must be blank (present, but without index sequences)
Note: A similar method applies when demultiplexing a run without the i7 index.
  • OverrideCycles with the N# for the i7. For example: OverrideCycles,Y151;N8;I8;Y151
  • Index column should be blank (present, but without sequences)
Note: If a user creates the sample sheet using Run Planning in BaseSpace, Run planning does not allow blank index sequence. As the workaround, input placeholder index sequences to match the index length, download the sample sheet, and remove the sequences from Index or Index2 column as needed.
For any feedback or questions regarding this article (Illumina Knowledge Article #5166), contact Illumina Technical Support [email protected].
Last modified 11d ago
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