Is there a maximum number of cycles for index reads on the NextSeq 1000/2000?

Background

The maximum number of cycles for an index read is dependent on the BCL convert version. The BCL Convert version is checked in both BaseSpace Run Planning and the local Sample Sheet mode.

• BCL Convert 4.2 or lower is limited to 27 index cycles used for demultiplexing. For example:

  • A run setup up with Index 1 = 20 cycles and Index 2 = 10 cycles would not be permitted as it exceeds the 27 cycle limit.

  • A run setup with Index 1 = 10 followed by a 10 bp UMI and Index 2 = 10 cycles would be allowed because only 20 cycles are used for demultiplexing.

• BCL Convert 4.3 and higher, the Index length has been increased to 27 cycles for each Index Read.

  • BCL Convert 4.3 is not currently compatible with the NextSeq 1000/2000 Control Software and the analysis would need to take on a separate local server or in BaseSpace.

There is NO maximum index read length within NextSeq 1000/2000 Control Software for the Manual Mode option as no Secondary Analysis is performed so it does not use BCL Convert.

NextSeq 1000/2000 Control Software displays a warning when Index 1 and Index 2 are fewer than 6 cycles. The warning will not display if Index 1 or Index 2 is 0 cycles.

Important Note: it is not possible to perform only an Index 2 read; to enable an Index 2 read, an Index 1 read must also be performed.

For further information, see Knowledge Base articleMaximum read length for Illumina sequencing platforms.

For any feedback or questions regarding this article (Illumina Knowledge Article #3459), contact Illumina Technical Support [e-mail].