A fast, integrated workflow for producing enriched and indexed sequencing libraries from a broad range of sample types and RNA input quantities.
Assay time
Hands-on time
Input quantity
Illumina RNA Prep with Enrichment uses on-bead tagmentation followed by a simplified, single 90-minute hybridization step to provide an exceptionally fast solution for RNA sequencing (RNA-Seq) applications.
Prepare and enrich sequencing-ready libraries in less than nine hours total time with less than two hours of hands-on time. Combine with an Illumina high-throughput sequencing instrument for significantly more samples per run.
Enable delivery of reproducible results from limited starting material and damaged or degraded samples, such as formalin-fixed, paraffin- embedded (FFPE) tissue sections.
Maximize discovery power at fraction of the sequencing depth by providing exceptional capture efficiency and coverage uniformity, minimizing required sequencing depth for accurate detection without bias.
Support discovery studies such as allele-specific expression, fusion detection, biomarker screening, and more using a variety of prebuilt, validated probes or custom v2 probe panels for maximum flexibility. Design custom probe panels using DesignStudio software.
| Assay time | < 9 hr |
|---|---|
| Automation capability | Liquid handling robot(s) |
| Content specifications | Captures custom targets with custom v2 Oligos designed in DesignStudio assay tool or entire coding transcriptome when used with the Illumina Exome Panel |
| Description | A reproducible, economical solution enabling targeted transcript detection and discovery of splice variants, gene fusions, and small nucleotide variants from a broad range of sample types and inputs including formalin-fixed, paraffin-embedded (FFPE) tissue sections and other low-quality samples |
| Hands-on time | < 2 hr |
| Input quantity | 10 ng RNA, 20 ng FFPE RNA, up to 1000 ng high-quality RNA |
| Instruments | NextSeq 550 System, NextSeq 2000 System, NextSeq 1000 System, NextSeq 550Dx in Research Mode, NovaSeq X System, NovaSeq 6000Dx in Research Mode, NovaSeq 6000 System, NovaSeq X Plus System, MiSeq i100 System, MiSeq i100 Plus System |
| Mechanism of action | Bead-linked transposome |
| Method | mRNA sequencing, Targeted RNA sequencing, Target enrichment |
| Multiplexing | Up to 384 Unique Dual Indexes (UDIs) |
| Nucleic acid type | RNA |
| Specialized sample types | Low-input samples, FFPE tissue |
| Species category | Any species, Human, Virus |
| Strand specificity | Non-stranded |
| Technology | Sequencing |
| Variant class | Single nucleotide polymorphisms (SNPs), Gene fusions, Transcript variants, Single nucleotide variants (SNVs), Insertions-deletions (indels) |
The complete RNA Prep with Enrichment workflow requires use of a library preparation kit, a probe panel, and index adapters (all items sold separately).
Optional components include:
Illumina RNA Fast Hyb Enrichment—Enrichment reagents only, for customers processing at a different plexity than recommended
Illumina RNA Prep, (L) Tagmentation—Pre-enrichment reagents only, for customers processing at a different plexity than recommended
Illumina RNA Prep with Enrichment enables flexibility in sample type and RNA input amounts in the preparation of targeted sequencing libraries for a wide range of RNA-Seq applications that support detection and discovery studies such as allele-specific expression, fusion detection, biomarker screening, and more.
Illumina RNA Prep with Enrichment
| Instrument | Recommended number of samples | Read length |
|---|---|---|
| NextSeq 550 System | Illumina RNA Prep with Enrichment + Exome Panel Mid output: 5 High output: 16 (based on 25M reads per sample) |
2 × 100 bp |
| NextSeq 1000 System, NextSeq 2000 System | Illumina RNA Prep with Enrichment + Exome Panel P2: 16 P3: 48 (based on 25M reads per sample) |
2 × 100 bp |
| NovaSeq 6000 System | Illumina RNA Prep with Enrichment + Exome Panel SP: 32 S1: 64 S2: 164 S4: 384 (based on 25M reads per sample) |
2 × 100 bp |
| MiSeq System | Illumina RNA Prep with Enrichment + Viral Surveillance Panel or Pan-Coronavirus Panel Up to 25 samples (based on 1M reads per sample) |
2 × 150 bp |
| NextSeq 550 System | Illumina RNA Prep with Enrichment + Viral Surveillance Panel or Pan-Coronavirus Panel Up to 384 samples (based on 1M reads per sample) |
2 × 150 bp |
Use NGS to discover novel microbes, monitor outbreaks, analyze food sources, and more.
Detect cancer gene expression and transcriptome changes and identify novel cancer transcripts with RNA-Seq for research.
| Illumina RNA Prep with Enrichment | TruSeq RNA Exome | Illumina Stranded mRNA Prep | |
|---|---|---|---|
| Assay time | < 9 hr | ~2 days | 6.5 hr |
| Automation capability | Liquid handling robot(s) | Liquid handling robot(s) | Liquid handling robot(s) |
| Content specifications | Captures custom targets with custom v2 Oligos designed in DesignStudio assay tool or entire coding transcriptome when used with the Illumina Exome Panel | Captures the coding transcriptome/RNA exome | Captures the coding transcriptome with strand information |
| Description | A reproducible, economical solution enabling targeted transcript detection and discovery of splice variants, gene fusions, and small nucleotide variants from a broad range of sample types and inputs including formalin-fixed, paraffin-embedded (FFPE) tissue sections and other low-quality samples | Gives researchers a clear, comprehensive view of the coding transcriptome with precise strand information and a reduced sample input requirement. | A simple, cost-effective solution for analysis of the coding transcriptome with precise strand information |
| Hands-on time | < 2 hr | ~11 hr | < 3 hr |
| Input quantity | 10 ng RNA, 20 ng FFPE RNA, up to 1000 ng high-quality RNA | 10 ng total RNA from fresh/frozen samples, or 20 ng total RNA from FFPE samples | 25-1000 ng standard-quality total RNA |
| Instruments | NextSeq 550 System, NextSeq 2000 System, NextSeq 1000 System, NextSeq 550Dx in Research Mode, NovaSeq X System, NovaSeq 6000Dx in Research Mode, NovaSeq 6000 System, NovaSeq X Plus System, MiSeq i100 System, MiSeq i100 Plus System | NextSeq 550 System, NextSeq 2000 System, NextSeq 1000 System, NovaSeq X System, NextSeq 500 System, NovaSeq 6000 System, NovaSeq X Plus System | NextSeq 550 System, NextSeq 2000 System, NextSeq 1000 System, NovaSeq X System, NextSeq 500 System, NovaSeq 6000 System, NovaSeq X Plus System |
| Mechanism of action | Bead-linked transposome | Biotinylated capture probes that target coding RNA. Does not require RNA with poly-A tails. | PolyA capture, ligation-based addition of adapters and indexes |
| Method | mRNA sequencing, Targeted RNA sequencing, Target enrichment | Exome sequencing, mRNA sequencing | mRNA sequencing |
| Multiplexing | Up to 384 Unique Dual Indexes (UDIs) | Up to 24 single, 96 combinatorial (CD) dual | Up to 384 Unique Dual Indexes (UDIs) |
| Nucleic acid type | RNA | RNA | RNA |
| Specialized sample types | Low-input samples, FFPE tissue | Low-input samples, FFPE tissue | Not FFPE-compatible, Low-input samples |
| Species category | Any species, Human, Virus | Human | Mammalian, Bovine, Mouse, Human, Rat |
| Strand specificity | Non-stranded | Stranded | Stranded |
| Technology | Sequencing | Sequencing | Sequencing |
| Variant class | Single nucleotide polymorphisms (SNPs), Gene fusions, Transcript variants, Single nucleotide variants (SNVs), Insertions-deletions (indels) | Single nucleotide polymorphisms (SNPs), Novel transcripts, Transcript variants | Single nucleotide polymorphisms (SNPs), Gene fusions, Novel transcripts, Transcript variants |
Library Prep & Array Kit Selector
Find the right sequencing library preparation kit or microarray for your needs. Filter by method, species, and more. Compare, share, and order kits.
On-bead tagmentation and a single, 90-min hybridization step combine to deliver a faster workflow with fewer steps compared to TruSeq RNA Exome.
Illumina RNA Prep with Enrichment achieves high data concordance between input amounts of 10 ng and 100 ng total RNA from universal human reference (UHR).
Libraries prepared from 10 ng RNA from the K-562 cell line using Illumina RNA Prep with Enrichment and the Illumina Exome Panel resulted in successful detection of the BCR-ABL1 gene fusion. Top alignment track shows all reads, bottom track shows only reads supporting the BCR-ABL1 fusion gene.
Libraries prepared from 10 ng UHR RNA and 20 ng FFPE RNA using Illumina RNA Prep with Enrichment and the Illumina Exome Panel show more than 85% of the data generated aligned to coding transcripts and UTRs. Data from TruSeq RNA Exome libraries are shown for comparison.
100% call rate for BCR-ABL1 and TPM3-NTRK1 gene fusions across six replicates of the K-562 cell line (RNA integrity number, RIN = 7.4, DV200 = 90%) and a colorectal cancer cell line (RIN = 2.5, DV200 = 85%).
| Fusion (source) | RIN | RNA input | Detection |
| BCR-ABL 1 (K-562) | 7.4 | 10 ng | 6/6 replicates (100%) |
| TPM3-NTRK1 (colorectal cancer) | 2.5 | 20 ng | 6/6 replicates (100%) |
This Illumina Technical Support webinar presents the Illumina RNA Prep with Enrichment workflow. This webinar is intended for users that are interested in targeted RNA sequencing using enrichment with fixed and custom panels.
RNA Sequencing: Introduction to Illumina RNA library preparation workflows
RNA Sequencing: Introduction to Illumina’s RNA library preparation workflows. This session reviews sample preparation kit options for RNA input.
RNA Sequencing: Best practices for Illumina RNA Prep protocols
This webinar covers best practices for bead handling, quantification methods, library QC, and troubleshooting view.
Advancements in translational biomarker identification from cell-free RNA
Liquid biopsy is most often associated with cell-free DNA, but new research has shown additional value in interrogating a broader diversity of analytes in the blood. Cell-free RNA (cfRNA) is one such molecule that has been shown to reflect dynamic changes in many tissues. Researchers at Illumina have optimized key variables to enable scalable, end-to-end workflow solutions for whole-transcriptome library preparations of cfRNA.
Comprehensive solution, from probe design to data analysis, for multiple targeted sequencing applications. Our custom-designed panels ensure high uniformity and coverage, with enhanced coverage of GC-rich and difficult to sequence areas.
Illumina® RNA Prep with Enrichment, (L) Tagmentation (16 Samples)
20040536
Includes reagents for preparing and enriching 16 libraries (16, 1-plex enrichment reactions). Purchase enrichment probe panel, Purification Beads and index adapters separately.
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Illumina® RNA Prep with Enrichment, (L) Tagmentation (96 Samples)
20040537
Includes reagents for preparing and enriching 96 libraries (32, 3-plex enrichment reactions). Purchase enrichment probe panel, Purification Beads and index adapters separately.
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Illumina® DNA/RNA UD Indexes Set A, Tagmentation (96 Indexes, 96 Samples)
20091654
Illumina® DNA/RNA UD Indexes Set A, Tagmentation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples. Purchase library prep and enrichment reagents and probe panels separately.
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Illumina® DNA/RNA UD Indexes Set B, Tagmentation (96 Indexes, 96 Samples)
20091656
Illumina® DNA/RNA UD Indexes Set B, Tagmentation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples. Purchase library prep and enrichment reagents and probe panels separately.
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Illumina® DNA/RNA UD Indexes Set C, Tagmentation (96 Indexes, 96 Samples)
20091658
Illumina® DNA/RNA UD Indexes Set C, Tagmentation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples. Purchase library prep and enrichment reagents and probe panels separately.
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Illumina® DNA/RNA UD Indexes Set D, Tagmentation (96 Indexes, 96 Samples)
20091660
Illumina® DNA/RNA UD Indexes Set D, Tagmentation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples. Purchase library prep and enrichment reagents and probe panels separately.
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Illumina Custom Enrichment Panel V2 (1536 µl, 120 bp)
20111339
Illumina Custom Enrichment Panel V2 enables 120 bp double-stranded custom panel design from 100 up to 1,000,000 probes using DesignStudio or Concierge Services and provides 1536 µl of material (sufficient for 384 enrichment reactions when paired with Illumina DNA Prep with Enrichment). Library prep, enrichment, and index adapter reagents need to be ordered separately.
Illumina Custom Enrichment Panel V2 (384 µl, 120 bp)
20073952
Illumina Custom Enrichment Panel V2 enables 120 bp double-stranded custom panel design from 100 up to 1,000,000 probes using DesignStudio or Concierge Services and provides 384 µl of material (sufficient for 96 enrichment reactions when paired with Illumina DNA Prep with Enrichment). Library prep, enrichment, and index adapter reagents need to be ordered separately.
Illumina Custom Enrichment Panel V2 (32 µl, 120 bp)
20073953
Illumina Custom Enrichment Panel V2 enables 120 bp double-stranded custom panel design from 100 up to 1,000,000 probes using DesignStudio or Concierge Services and provides 32 µl of material (sufficient for 8 enrichment reactions when paired with Illumina DNA Prep with Enrichment). Library prep, enrichment, and index adapter reagents need to be ordered separately.
Illumina Exome Panel – Enrichment Oligos Only
20020183
Sufficient for 8 enrichment reactions when paired with Illumina DNA Prep with Enrichment, 12 reactions when paired with TruSeq RNA Exome, and 32 reactions when paired with Illumina RNA prep with enrichment. Purchase library prep and enrichment reagents and index adapters separately.
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Illumina RNA Prep with Enrichment – Customer Site
20044763
2 day on-site training for the Illumina RNA Prep with Enrichment Kit. Hands-on training includes quantification of RNA input as well as best practices, troubleshooting tips, and training on Illumina-supported analysis tools specific to the application workflow.
Illumina RNA Fast Hyb Enrichment (16 Samples)
20040540
Includes fast hybridization enrichment reagents from the Illumina RNA Prep with Enrichment kit for 16, 1-plex enrichment reactions. Supports customers processing at a different plexity from what is recommended. Pre-enrichment reagents not included. For complete prep with enrichment order the Illumina RNA Prep with Enrichment kit.
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Illumina RNA Prep, (L) Tagmentation (16 Samples)
20040542
Includes pre-enrichment module from the Illumina RNA Prep with Enrichment kit for 16 libraries. Supports customers processing at a different plexity from what is recommended. Enrichment reagents and Purification Beads not included. For complete prep with enrichment order the Illumina RNA Prep with Enrichment kit.
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Illumina recommends using Agilent Technologies Universal Human Reference RNA (UHR) (Catalog no. 740000) as a positive control sample.
The Illumina Exome Panel and Illumina Custom Enrichment Panel v2 can be used with the Illumina RNA Prep and Enrichment kit.
eBLTL are enrichment bead-linked transposomes that tagment cDNA by fragmenting and adding adapter tag sequences in a single and rapid reaction step.
The 16-reaction kit enables a 1-plex workflow while the 96-reaction kit enables a 3-plex workflow. Additional accessory products can be added to either workflow to process at a different plexity from what is recommended.
Targeted custom enrichment panels, providing simple and efficient solutions for multiple targeted sequencing applications.
Reagent kits for the NovaSeq X Series include a reagent cartridge, buffer cartridge, flow cell, lyo insert, pre-load buffer, and library tube strip.
Reagents for the MiSeq i100 Series feature easy-to-use cartridges and multiple flow cell configurations for flexible sequencing options.
A rapid, cost-effective workflow for accurate, unbiased detection of the protein-coding transcriptome with precise measurement of strand information.
Reagents for the NextSeq 1000/2000 System feature easy-to-use cartridges and multiple flow cell configurations for flexible sequencing options.
Fast, simple library prep workflow for RNA exome analysis, combining cDNA synthesis and adapter tagging into a single step.
Accessible and highly scalable single-cell RNA-Seq solution for mRNA capture, barcoding, and library prep without complex workflows or microfluidics
Reagent v2.5 kits use a resilient flow cell that maintains starting fluorescent intensity, improving stability and performance compared to earlier versions.
Library prep and enrichment kit that delivers highly sensitive, comprehensive respiratory pathogen identification and antimicrobial resistance insights.
Looking for more? Reach out for information about our products and services, answers to questions about our technology.
