NGS Barcode Adapters for Multiplexed Sequencing
NEXTFLEX™ barcode adapters are available in multiple UDI index lengths and UDI-UMI formats for Illumina® and Element® sequencing workflows. Designed for use with NEXTFLEX and other TruSeq™-style (ligation-based) library prep workflows, these adapters support routine multiplexing, high-throughput indexing, and molecular-level read tracking.
Need help choosing an indexing strategy?
Our team can help you select the right NEXTFLEX™ barcode adapter format based on your library prep workflow, sequencing platform, multiplexing scale, and UMI requirements.
Selection guide
| Indexing need | Revvity solution | Details |
|---|---|---|
| Multiplex samples with unique dual indexes | NEXTFLEX™ Unique Dual Index Barcodes (8NT- 384) | UDI adapters for NEXTFLEX™ and other TruSeq-style ligation-based library prep workflows. Available in multiple plate configurations to support routine multiplexing. |
| Expand indexing capacity for high-throughput studies | NEXTFLEX™ Unique Dual Index Barcodes (10NT – 1,536) | Expanded UDI options for high-throughput sequencing projects that require higher multiplexing capacity. |
| Add molecule-level read tracking | NEXTFLEX™ UDI-UMI Barcodes | Combines unique dual indexes with UMIs for workflows involving duplicate identification, molecular counting, or consensus analysis. |
Need blockers for target enrichment?
Explore NEXTFLEX™ Universal Blockers for hybrid capture workflows using indexed or UMI-containing libraries.
Figure 1: Schematic showing a library insert flanked by adapter sequences used in NEXTFLEX® UDI and UDI-UMI barcode adapters. P5 and P7 indicate flow cell binding sites for Illumina® sequencing workflows, SP1 and SP2 indicate sequencing primer binding sites, i5 and i7 indicate unique dual indexes used for sample identification, and UMI indicates the 9 bp unique molecular identifier used to tag individual molecules within a library.
Key considerations when choosing NGS barcode adapters
Barcode adapter selection depends on more than the number of samples being sequenced. Index design, multiplexing scale, UMI requirements, and library prep compatibility can all affect how libraries are pooled, sequenced, and assigned during analysis.
Multiplexing scale
UDI adapters provide unique i5 and i7 index combinations for sample assignment across pooled libraries. Larger studies may benefit from expanded UDI sets with 10 nt indexes, which support higher multiplexing capacity for high-throughput sequencing projects.
Index design
Well-designed UDI sets do more than identify pooled libraries. Unique i5/i7 index pairs help reduce sample misassignment from index hopping, while sufficient Hamming distance helps protect against index read errors during demultiplexing. Balanced base composition across index reads also supports reliable base calling in multiplexed sequencing runs.
UMI-based molecular tracking
UDI-UMI adapters combine sample indexing with unique molecular identifiers, allowing reads derived from the same original molecule to be grouped during analysis. This can support applications involving duplicate identification, molecular counting, or consensus analysis.
Schematic showing a library insert flanked by adapter sequences used in NEXTFLEX® UDI and UDI-UMI barcode adapters. P5 and P7 indicate flow cell binding sites for Illumina® sequencing workflows, SP1 and SP2 indicate sequencing primer binding sites, i5 and i7 indicate unique dual indexes used for sample identification, and UMI indicates the 9 bp unique molecular identifier used to tag individual molecules within a library.
Library prep compatibility
Adapter choice should be compatible with the library prep chemistry used in your workflow. NEXTFLEX™ barcode adapters are designed for use with NEXTFLEX™ and other TruSeq-style ligation-based workflows, allowing labs to evaluate new indexing options without having to change established library prep processes.
For research use only. Not for use in diagnostic procedures.
NEXTFLEX™ barcode adapter products
Explore NEXTFLEX™ adapter options for UDI-based sample indexing, expanded multiplexing capacity, and UMI-enabled molecular tracking in compatible NGS workflows.
NEXTFLEX™ Unique Dual Index Barcodes
NEXTFLEX™ Unique Dual Index Barcodes are available in multiple UDI configurations for multiplexed sequencing workflows, including expanded 10 nt options for higher-throughput projects. These adapters are a good fit for labs that need reliable sample indexing while maintaining compatibility with NEXTFLEX™ or other TruSeq-style ligation-based library prep workflows.
NEXTFLEX™ UDI-UMI Barcodes
NEXTFLEX™ UDI-UMI Barcodes combine unique dual indexing with integrated UMIs for workflows that require both sample identification and molecule-level tracking. These adapters are best suited for applications where UMI information will be used downstream for duplicate identification, molecular counting, or consensus analysis.
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FAQs
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What is the difference between UDI and UMI adapters?
Unique dual index adapters use distinct i5 and i7 index sequences to identify each sample in a multiplexed sequencing run. UMI adapters include unique molecular identifiers in addition to sample indexes, allowing reads derived from the same original molecule to be grouped during analysis.
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When should I use UDI-UMI adapters instead of UDI adapters?
Use UDI-UMI adapters when your workflow requires both sample indexing and molecule-level tracking. UMI information can support applications involving duplicate identification, molecular counting, and consensus analysis. If your primary need is sample multiplexing, UDI adapters are usually the more direct option.
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How do unique dual indexes help reduce index hopping?
Unique dual indexes assign each library a distinct i5/i7 index pair. This helps reduce sample misassignment from index hopping because an incorrectly assigned read is less likely to match a valid expected index pair. Well-designed index sets with sufficient spacing between sequences also help protect against index read errors during demultiplexing.
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What is the difference between 8 nt and 10 nt UDI adapters?
Both 8 nt and 10 nt UDI adapters support unique dual indexing for multiplexed NGS workflows. The 10 nt UDI format provides expanded indexing capacity, making it useful for high-throughput studies that require more unique index combinations across larger sequencing projects.
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Can NEXTFLEX™ barcode adapters be used with non-NEXTFLEX library prep kits?
NEXTFLEX™ barcode adapters are designed for use with NEXTFLEX™ and other TruSeq-style ligation-based library prep workflows. This allows labs to evaluate new indexing options without replacing an established library prep process. They are compatible with Illumina® and Element® sequencing workflows when used with appropriate library prep chemistry, run setup, sample sheet configuration, and demultiplexing settings.
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Do I need Universal Blockers with these barcode adapters?
Universal Blockers are used for hybrid capture or target enrichment workflows where adapter, index, or UMI sequences need to be blocked during probe hybridization. They are not required for every NGS workflow. For standard sequencing workflows without hybrid capture, barcode adapters can be selected independently of blockers.
