Targeted RNA Sequencing
What Is Targeted RNA Sequencing?
Targeted RNA sequencing is a precision‐focused NGS approach that enriches and quantifies only the transcripts you care about—rather than sequencing the entire transcriptome—by using either PCR primers (amplicon panels) or hybridization probes tiled across specific genes, exons, splice junctions, or fusion breakpoints. In an amplicon‐based workflow, multiplexed gene‐specific primers amplify dozens to hundreds of targets in a single reaction, delivering ultra‐deep coverage (>1,000×) with minimal input (as little as 5 ng RNA) and a streamlined, 1–2 day library prep. The hybrid‐capture workflow uses biotinylated oligonucleotide probes that selectively bind target cDNAs—enabling uniform enrichment of larger panels (up to several megabases of cumulative target space) and robust performance across GC‐rich or repetitive regions, UTRs, and noncoding RNAs. Both approaches incorporate adapter ligation, dual‐index barcoding, and optional Unique Molecular Identifiers (UMIs) to collapse PCR duplicates and correct amplification bias. After enrichment, libraries are sequenced (typically 1–5 million reads per sample) on Illumina platforms, and data are aligned to a custom reference, producing digital counts for each target, fusion calls, and splice‐variant quantification. This highly efficient design dramatically reduces sequencing costs and data complexity—while delivering the sensitivity, reproducibility, and scalability needed for large‐cohort studies, clinical assays, and longitudinal biomarker monitoring.
Advantages of Targeted RNA Sequencing
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Ultra-High Sensitivity: Achieve >1,000× coverage on your selected genes to reliably quantify rare transcripts, splice variants, and fusion junctions.
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Cost & Data Efficiency: Use just 1–5 million reads per sample—saving up to 90% on sequencing and storage compared to whole-transcriptome assays.
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Fully Customizable Panels: Design assays for 5–500+ targets, including exons, UTRs, fusion breakpoints, or noncoding RNAs without revalidating a whole-genome workflow.
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Dual Workflow Options:
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Amplicon: Rapid 1–2 day prep with ≥5 ng input, ideal for small panels and degraded FFPE.
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Hybrid Capture: Uniform enrichment of large panels (up to 2 Mb) across GC-rich or repetitive regions.
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Robust on Challenging Inputs: Reliable performance on DV200 ≥ 30% RNA, FFPE samples, biofluids, or as little as 5 ng total RNA.
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Digital Quantitation with UMIs: Optional barcodes collapse PCR duplicates for true molecule counts and confidence in low-frequency calls.
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Rapid Turnaround & Scalability: Standard delivery in 7–10 days (rush in 3–5 days); multiplex 96–384 samples per run.
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Actionable Insights & Reporting: End-to-end support from panel design to interactive reports with digital counts, fusion/isoform calls, and pathway enrichment.
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Pathway-Focused Studies: Immune profiling (cytokines, checkpoints), oncology panels (oncogenes, tumor suppressors), and metabolic or signaling pathways.
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Validation of Discovery Hits: Rapidly confirm differentially expressed genes from discovery RNA-Seq or microarray screens.
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Biomarker Panels: High-throughput screening of clinical cohorts for diagnostic or prognostic markers.
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Fusion & Splice Variant Detection: Detect clinically relevant gene fusions, exon skipping events, and alternative transcript isoforms.
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Degraded & FFPE Samples: Reliable quantitation in archival specimens or cytology slides.
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Drug Response & Toxicity Assays: Monitor gene expression changes in preclinical models or patient samples.
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Time-Course & Perturbation Experiments: Quantitatively track transcript dynamics across treatments, doses, or time points.
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Efficient Disease Monitoring: Track disease progression and therapeutic response with high-resolution expression profiling.
What is Targeted RNA Sequencing Used For?
Targeted RNA Sequencing with AUGenomics
Sample Submission
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Accepted Sample Types: Total RNA from tissues, cells, FFPE blocks, or biofluids
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Input Requirement: 10–100 ng of total RNA (≥5 ng supported for some panels)
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Sequencing Recommendations:
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Amplicon or capture-based enrichment workflows
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Read depth: 1–5 million reads/sample depending on panel size
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Multiplexing up to hundreds of samples per run
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Please refer to our Shipping Guidelines for project-specific guidance.
Turnaround Time
Typical turnaround is 7-10 business days from sample receipt. Expedited options are available depending on project scope and sequencing depth.
Frequently Asked Questions (FAQs)
Q: What’s the difference between targeted RNA-seq and mRNA-seq?
A: Targeted RNA-seq focuses on a defined panel of genes, offering greater efficiency, lower cost, and higher coverage for those genes. mRNA-seq captures the entire protein-coding transcriptome.
Q: Can I use targeted RNA-seq for FFPE samples?
A: Yes. Our workflows are optimized for fragmented or degraded RNA and are frequently used with FFPE-derived material.
Q: How do I choose between amplicon and capture?
A: Use amplicon for small panels (<100 kb), rapid prep, and low-input/FFPE; choose capture for larger panels, complex loci, or when capturing flanking regions and UTRs.
Q: What read depth do I need?
A: Typically 1–5 M reads per sample yields >1,000× coverage on targets; panel size and number of targets will drive specific recommendations.
Got more questions? Contact our team and get a free consultation anytime. info@augenomics.com
Glossary of Terms
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Targeted RNA-seq: Sequencing method focusing on specific RNA transcripts or genes of interest
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Amplicon-Based Sequencing: PCR-based enrichment of target RNA regions
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Hybrid Capture: Probe-based enrichment method for selected transcripts
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Multiplexing: Sequencing multiple samples or targets in a single run
