Overview
Illumina Stranded mRNA Prep provides a streamlined RNA sequencing (RNA-Seq) solution for clear and comprehensive analysis across the coding transcriptome for discovery of features such as novel isoforms, gene fusions, and allele-specific expression.
Key features
Precise measurement of strand orientation— Enables detection of antisense transcription, enhances transcript annotation, and increases alignment efficiency
Exceptional performance— Provides excellent polyA capture efficiency and coverage uniformity, minimizing required sequencing depth for accurate, unbiased detection of the coding transcriptome. Learn more about the technology
Exceptional performance— Provides excellent polyA capture efficiency and coverage uniformity, minimizing required sequencing depth for accurate, unbiased detection of the coding transcriptome.
Learn more about the technology
High-quality data from low-input samples— Achieves highly sensitive results from as little as 25 ng of high-quality RNA*
Fast library preparation workflow— Prepares libraries ~40% faster than TruSeq Stranded mRNA; shorter incubation times and reduced sample cleanup steps result in a total assay time of 6.5 hr
*Not compatible with FFPE samples
Applications
Illumina Stranded mRNA Prep enables preparation of sequencing-ready transcriptome libraries in a single day with low sample inputs of standard (non-degraded) RNA from a wide range of sample types.
Example workflow
Illumina Stranded mRNA Prep
NextSeq 550 System
NextSeq 1000/2000 Systems
NovaSeq 6000 System
DRAGEN RNA
RNA-Seq Differential Expression
RNA-Seq Alignment App
Project recommendations
Instrument: Recommended number of samples
NextSeq 550 System: Mid output: 5 High output: 16 (based on 25M reads per sample)
Mid output: 5 High output: 16 (based on 25M reads per sample)
2 × 75 bp
NextSeq 2000 System: P2: 16 P3: 48 (based on 25M reads per sample)
P2: 16 P3: 48 (based on 25M reads per sample)
NovaSeq 6000 System: SP: 32 S1: 64 S2: 164 S4: 384 (based on 25M reads per sample)
SP: 32 S1: 64 S2: 164 S4: 384 (based on 25M reads per sample)
Related applications and methods
mRNA sequencing
mRNA sequencing (mRNA-Seq) using NGS can identify both known and novel transcripts, and measure transcript abundance.
Gene expression analysis
Learn how to analyze transcriptome changes or profile genome-wide gene expression levels in a single experiment with next-generation sequencing methods.
Comparison of performance metrics
(Left) Performance metrics for Illumina Stranded mRNA Prep. (Right) Illumina Stranded mRNA Prep was compared against TruSeq Stranded mRNA. Illumina Stranded mRNA Prep showed superior performance, particularly with an input of 25 ng UHR RNA.
Greater gene discovery at low input
Illumina Stranded mRNA Prep enables greater gene detection with low RNA inputs, as compared to TruSeq Stranded mRNA. The number of genes detected is reported at 30M subsampled paired-end reads PF. More genes detected at 1× coverage is an indicator of greater sensitivity.
High concordance with legacy kit
Illumina Stranded mRNA Prep produces highly concordant data with TruSeq Stranded mRNA at (A) equivalent inputs of 100 ng UHR RNA and (B) with reduced input of 25 ng vs 100 ng UHR RNA.
High data concordance
Illumina Stranded mRNA Prep achieves high data concordance between (A) technical replicates of 25 ng UHR RNA and (B) between input amounts of 25 ng and 100 ng UHR RNA. Libraries were sequenced at 2 × 74 bp, subsampled to 30M reads. Data analysis was performed using the BaseSpace RNA-Seq Alignment App.
Illumina Stranded mRNA Prep workflow
Illumina Stranded mRNA Prep delivers a fast workflow with reduced hands-on time. Times may vary depending on equipment used, number of samples processed, automation procedures, or user experience.
What is stranded mRNA-Seq?
To accurately determine gene expression from overlapping genes, it is important to retain the information found on the strand of origin. Stranded RNA-Seq allows the first and second cDNA strands to be distinguished so that the second strand can be degraded while the first cDNA strand (strand of origin) will undergo further PCR amplification.
How long does Illumina Stranded mRNA library prep take?
Library preparation takes 6.5 hr with less than 3 hr of hands-on time.
How do you prepare mRNA for sequencing with Illumina Stranded mRNA Prep?
Oligo(dT) magnetic beads capture and purify the mRNA containing polyA tails. The purified mRNA is fragmented and copied into first-strand cDNA using reverse transcriptase and random primers. In a second-strand cDNA synthesis step, dUTP replaces dTTP to achieve strand specificity. The final steps add adenine and thymine bases to fragment ends and ligate adapters. The resulting products are purified and selectively amplified for sequencing.
Illumina® Stranded mRNA Prep, Ligation (16 Samples).
20040532
A simple, scalable, cost-effective, rapid single-day solution for analyzing the coding transcriptome leveraging as little as 25 ng of total RNA input. Kit contains library prep reagents for 16 samples. Indexes sold separately, compatible only with Illumina RNA UD indexes, Ligation View less
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Illumina® Stranded mRNA Prep, Ligation (96 Samples).
20040534
A simple, scalable, cost-effective, rapid single-day solution for analyzing the coding transcriptome leveraging as little as 25 ng of total RNA input. Kit contains library prep reagents for 96 samples. Indexes sold separately, compatible only with Illumina RNA UD indexes, Ligation. View less
Illumina® RNA UD Indexes Set A, Ligation (96 Indexes, 96 Samples)
20091655
Illumina® RNA UD Indexes Set A, Ligation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples and RNA Index Anchor plate. Used with Illumina Stranded Total RNA Prep and Illumina Stranded mRNA Prep Kits. Purchase library prep reagents separately. View less
Illumina® RNA UD Indexes Set B, Ligation (96 Indexes, 96 Samples)
20091657
Illumina® RNA UD Indexes Set B, Ligation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples and RNA Index Anchor plate. Used with Illumina Stranded Total RNA Prep and Illumina Stranded mRNA Prep Kits. Purchase library prep reagents separately. View less
Illumina® RNA UD Indexes Set C, Ligation (96 Indexes, 96 Samples)
20091659
Illumina® RNA UD Indexes Set C, Ligation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples and RNA Index Anchor plate. Used with Illumina Stranded Total RNA Prep and Illumina Stranded mRNA Prep Kits. Purchase library prep reagents separately. View less
Illumina® RNA UD Indexes Set D, Ligation (96 Indexes, 96 Samples)
20091661
Illumina® RNA UD Indexes Set D, Ligation (96 Indexes, 96 Samples) Includes 96, 10 bp indexes sufficient for labeling 96 samples and RNA Index Anchor plate. Used with Illumina Stranded Total RNA Prep and Illumina Stranded mRNA Prep Kits. Purchase library prep reagents separately. View less
Illumina Stranded mRNA LP Kit Training - Customer Site
20044765
2-day on-site training for the Illumina Stranded mRNA Library Preparation Kit. Hands-on training includes quantification of mRNA input as well as best practices, troubleshooting tips, and training on Illumina-supported analysis tools specific to the application workflow. View less
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