Introduction to mRNA Sequencing
mRNA sequencing (mRNA-Seq) has rapidly become the method of choice for analyzing the transcriptomes of disease states, of biological processes, and across a wide range of study designs. In addition to being a highly sensitive and accurate means of quantifying gene expression, mRNA-Seq can identify both known and novel transcript isoforms, gene fusions, and other features as well as allele-specific expression. mRNA-Seq delivers a complete view of the coding transcriptome that is not restricted by the filter of prior knowledge.
mRNA-Seq in 3 Simple Steps
Analyze the coding transcriptome in standard mRNA samples with this seamless workflow solution.
Advantages of mRNA Sequencing
mRNA-Seq provides a number of advantages over gene expression arrays in analyzing the transcriptome.
- Offers a broader dynamic range, enabling more sensitive and accurate measurement of fold changes in gene expression
- Captures both known and novel features
- Can be applied across a wide range of species
Transitioning from Arrays to mRNA-Seq
Expression Analysis developed tools to make it easier to compare mRNA-Seq results with previous array data.
Accurate, High-Resolution View of the Transcriptome
Ratio compression is an established technical limitation of gene expression arrays that reduces dynamic range and can mask or alter measured transcriptional changes.1–3 In contrast, mRNA-Seq is not subject to this bias and provides more comprehensive and accurate measurements of gene expression changes.
Additionally, mRNA-Seq can provide strand information, which enables the detection of antisense expression, allows more accurate quantification of overlapping transcripts, and increases the percentage of alignable reads.
Autism and mRNA-Seq
Stanley Lapidus, President, CEO, and Founder, SynapDx, discusses how the company is using mRNA-Seq to study autism.
Recommended mRNA-Seq Workflow for Standard Samples
Library Prep
TruSeq Stranded mRNA Library Prep Kit
Streamlined, cost-efficient, and scalable library preparation for mRNA-Seq.
Sequencing
NextSeq 550 System
Desktop sequencer supporting multiple applications, enabling sequencing of 5-16 mRNA samples in a single run.
Alignment Distribution Graph
Transcript Assembly Table
Differential Expression Graph
Comprehensive mRNA-Seq Workflow
Illumina sequencing by synthesis (SBS) chemistry is the most widely adopted NGS technology, producing approximately 90% of global sequencing data.*
In addition to our industry-leading data quality, Illumina offers integrated mRNA-Seq workflows that simplify the entire process, from library preparation to data analysis and biological interpretation.
Click on the below to view products for each workflow step.
TruSeq Stranded mRNA Library Prep Kit
Streamlined, cost-efficient, and scalable library preparation for mRNA-Seq, with precise measurement of strand orientation. Accurately measure gene and transcript abundance and detect both known and novel features in the coding transcriptome.
TruSeq RNA ExomeLow all-in cost solution for mRNA-seq of low-quality/FFPE samples. Accurately measure gene and transcript abundance and detect both known and novel features in the coding transcriptome.
Find the Right Library Prep Kit
Use this tool to determine the best kit for your needs.
MiSeq System
Speed and simplicity for focused applications, sequencing 1 mRNA sample per run.
NextSeq 550 SystemFlexible desktop sequencer supporting multiple applications, enabling 5–16 mRNA samples to be sequenced in a single run.
HiSeq 4000 SystemHigh-throughput RNA-Seq with up to 100 human transcriptomes on the HiSeq 4000 System.
NovaSeq 6000 System
Scalable throughput and flexibility for virtually any genome, sequencing method, and scale of project.
Platform Comparison ToolCompare sequencing platforms and identify the best system for your lab and applications.
Sequencing ReagentsFind kits that include sequencing reagents, flow cells, and/or buffers tailored to each Illumina sequencing system.
RNA Express App
Aligns RNA-Seq reads with the STAR aligner and assigns aligned reads to genes, followed by differential expression with DESeq2.
TopHat Alignment AppMaps reads, performs abundance estimations of reference genes and transcripts, calls variants, and offers optional fusion calling.
Cufflinks Assembly & Differential Expression (DE) AppAssembles novel transcripts and performs differential expression of novel and reference transcripts.
Integrative Genomics ViewerA genome browser developed by the Broad Institute of MIT and Harvard that displays NGS data for complex variant analysis.
Illumina DRAGEN Bio-IT PlatformThe Illumina DRAGEN (Dynamic Read Analysis for GENomics) Bio-IT Platform provides ultra-rapid secondary analysis of NGS data. A variety of apps are available, including one designed to enable gene fusion studies.
Genomatix Pathway System (GePS)
Associates single gene or list of genes with annotation data for pathways, diseases, tissues, and small molecules.
iPathway GuideDifferential gene expression, drug interaction, and disease analysis.
BaseSpace Sequence HubThe Illumina genomics computing environment for NGS data analysis and management.
BaseSpace Correlation EngineA growing library of curated genomic data to support researchers in identifying disease mechanisms, drug targets, and biomarkers.
Related Solutions
mRNA-Seq for Cancer Research
Monitoring gene expression changes with mRNA-Seq can help researchers identify biomarkers predictive of disease prognosis or response to therapy. Learn more about cancer RNA-Seq.
Gene Expression Analysis for Complex Disease Studies
RNA-Seq-based gene expression profiling studies can provide visibility into how genetic and environmental factors contribute to a broad range of complex diseases. Learn more about gene expression profiling.
RNA Drug Response Biomarker Discovery
Find out how to utilize RNA-Seq to identify novel RNA-based drug response biomarkers. Access resources designed to help new users adopt this application. Learn more about drug response RNA biomarker analysis.
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Additional Resources
Methods Guide
All the information you need, from BeadChips to library prep to sequencer selection and analysis. Use this guide to select the best tools for your lab.
Single-Cell mRNA-Seq
Dr. Norma Neff discusses how researchers at Stanford University are using single-cell mRNA-Seq to understand early development.
RNA-Seq Data Analysis
User-friendly software tools simplify mRNA-Seq data analysis for biologists, regardless of bioinformatics experience.
RNA-Seq for Gene Expression Studies
Illumina offers an integrated mRNA-Seq workflow for a deeper understanding of biology.
RNA-Seq of Low-Quality and FFPE Samples
RNA-Seq of formalin-fixed, paraffin-embedded (FFPE) and other low-quality samples offers valuable insights for disease research.
Paired-End RNA Sequencing
Paired-end RNA-Seq enables discovery applications such as detecting gene fusions and characterizing novel splice isoforms.
Core Sequencing Lab Benefits from BaseSpace Cloud
BRC-Seq uses BaseSpace for sample tracking as well as data analysis, management, and sharing.
Gene Panel and Array Finder
Identify sequencing panels or microarrays that target your genes of interest.
References
- Shi L, Tong W, Su Z, et al. Microarray scanner calibration curves: characteristics and implications. BMC Bioinformatics. 2005;6 Suppl 2:S11.
- Naef F, Socci ND, Magnasco M. A study of accuracy and precisions in oligonucleotide arrays: extracting more signal at large concentrations. Bioinformatics. 2003;19:178-184.
- Yuen T, Wurmbach E, Pfeffer RL, Ebersole BJ, Sealfon SC. Accuracy and calibration of commercial oligonucleotide and custom cDNA microarrays. Nucleic Acids Res. 2002;30:e48.
*Data calculations on file. Illumina, Inc., 2015