IDT for Illumina DNA/RNA UD Indexes FAQs

Unique dual indexes allow the index hopped reads to be bioinformatically removed. The ligation-based unique dual indexes have been released for TruSeq DNA PCR Free, TruSeq DNA Nano, TruSeq DNA Exome, TruSeq Stranded mRNA, and TruSeq Stranded Total RNA library prep kits. The IDT for Illumina DNA/RNA UD Indexes are extension based indexes that are compatible with the Illumina DNA Prep, Nextera XT, and Illumina DNA Prep with Enrichment library prep kits.

Note: When using with Nextera XT, you need to perform a manual normalization workflow instead of the bead-based normalization workflow with the IDT for Illumina DNA/RNA UD Indexes. The IDT for Illumina DNA/RNA UD Indexes are not compatible with bead-based normalization.

The level of index hopping will not change with IDT for Illumina DNA/RNA UD Indexes, but the index hopped reads can be removed bioinformatically from the demultiplexed data.

IDT for Illumina DNA/RNA UD Indexes are compatible with all of the Illumina sequencing platforms except HiSeq X.

See the setup page for your sequencing instrument.

No, the IDT for Illumina DNA/RNA UD Indexes are not compatible with the Nextera XT bead based normalization. Continue to use the Nextera XT Indexes if you want to use the bead-based normalization workflow.

They are not designed to be used with any other Illumina index sets.  It is not recommended to sequence libraries made from different index sets (i.e. Nextera CD Indexes, IDT for Illumina TruSeq UD Indexes, or Nextera XT Index Kits).

You can run different libraries from different indexes but they must be loaded onto different lanes.  It is not recommended to mix libraries of different lengths and from different index kits.

Mixing libraries made from different index kits are not recommended. The IDT for Illumina DNA/RNA UD Indexes have 10 base pair codes. If you want to run different libraries made from different index kits, load the different libraries from different index kits in different lanes. Mixing libraries from different index kits may compromise your results.

Yes, there are combinations for 2-plex to 96-plex within the plate. For more information, see the Index Adapter Pooling Guide.

The reagents are shipped on dry ice and must be stored at -25°C to -15°C upon arrival.

We have tested up to 24 freeze/thaws of frozen index adapters without observing any loss of activity.

There are 10 μl of dual index in each well with some overage to account for storage and automated workflows. 

Combining libraries from different index sets within the same lane is NOT supported, regardless of the index lengths, unless the Illumina recommendation specifies that the index sets can be used together (e.g. UDP set A with UDP set B). This is because index sets may have indexes that are not compatible with each other across the two sets.

These indexes are compatible with Nextera XT (manual normalization only, not compatible with bead-based normalization), Illumina DNA Prep, and Illumina DNA Prep with Enrichment.

No, there is no change to the index volume required for the compatible library prep.

For Illumina DNA Prep and Nextera XT, use 10 μl from each well. 

Libraries with the IDT for Illumina DNA/RNA UD Indexes are expected to migrate similarly to the libraries prepared with the CD indexes and the Illumina DNA Prep kit.

For more information, contact Illumina Technical Support.