Ion Ampliseq™

Targeted, quantitative gene expression analysis

Researchers employ targeted sequencing as a rapid and cost-effective alternative to whole transcriptome sequencing by focusing on selected sets of genes or genomic regions. Ion AmpliSeq™ technology delivers outstanding data quality with a simple and fast workflow for affordable analysis of human gene expression.

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Targeted Ion AmpliSeq™ RNA Panels enable scientists to choose from over 20,000 genes to survey gene expression fold changes starting with as little as 500 pg of unfixed RNA or 5 ng of formalin-fixed, paraffin-embedded (FFPE) RNA. The Ion PGM™ or Ion S5™ systems, coupled with Ion AmpliSeq RNA Panels, Torrent Suite™ Software, and Partek™ Flow™ Software, facilitates a unique and complete workflow solution.

Researchers employ targeted sequencing as a rapid and cost-effective alternative to whole transcriptome sequencing by focusing on selected sets of genes or genomic regions. The ability to efficiently and uniformly amplify just the RNA sequences of interest while keeping the RNA input amount can be a critical factor, especially in human disease research applications where minimal sample is commonly available. Targeted RNA sequencing is ideal for gene regulation analysis, correlating gene expression with phenotypic information, and cellular pathway analyses. Using Ion AmpliSeq technology, targeted RNA sequencing is a sequencing-based gene expression profiling method that allows quantification of messenger RNA (mRNA) or targeted non-coding RNA (ncRNA) levels for up to 300 customer-defined genes in a single amplification reaction. Importantly, the Ion AmpliSeq targeted RNA workflow utilizes very low amounts of input total RNA: 500 pg of unfixed RNA or 5 ng of RNA isolated from FFPE samples. Further, targeted RNA sequencing can be used to detect fusion transcripts from chromosomal rearrangements that are critical initiating steps of tumorigenesis. This technology, coupled with the Ion PGM or Ion S5 systems, Torrent Suite Software, and the Coverage Analysis plugin, enables researchers to go from precious FFPE samples to quantitative gene expression profiling in less than 24 hours.


Video: Dr. Audrey Papp from Ohio State University discusses targeted RNA sequencing using Ion AmpliSeq technology.

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Design your own Ion AmpliSeq RNA custom panel

Creating and ordering custom panels is easy with Ion AmpliSeq™ Designer—a free, online assay design tool that connects you directly to our assay design pipeline. Create and order custom RNA panels by simply selecting your targets of interest or entering your specific genomic content online, and we will design and create a customized Ion AmpliSeq™ panel just for you. Leveraging more than a decade of expertise powering the Applied Biosystems™ TaqMan® Custom Assay design pipeline, the Ion AmpliSeq Designer tool produces optimized primer designs in as little as a few minutes. You now have the flexibility to analyze panels of genes of your choosing, such as those implicated in a particular disease state or representing specific biological pathways with anywhere from 12 to 1,200 human amplicons in a single reaction. Each design accounts for GC content, is filtered for repeats and SNPs, and selects target exons that represent the maximum number of transcripts for genes having multiple transcript isoforms documented in standard databases such as RefSeq.

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Ready-to-use panels

In addition to custom panels, we offer the following Ion AmpliSeq RNA ready-to-use panels:

Ion AmpliSeq Transcriptome Human Gene Expression Kit

The Ion AmpliSeq™ Transcriptome Human Gene Expression Kit is a comprehensive tool for gene-level expression analysis. Targeting >20,000 human RefSeq transcripts in a single reaction with an FFPE-compatible workflow, the complete Ion AmpliSeq™ Transcriptome solution is compatible with existing downstream microarray or next-generation sequencing (NGS) analysis pipelines for fast and straightforward data interpretation.

Learn more about the Ion AmpliSeq Transcriptome Human Gene Expression Panel ›

Ion AmpliSeq RNA Fusion Lung Cancer Research Panel

Developed with input from the OncoNetwork Consortium, the Ion AmpliSeq™ RNA Fusion Lung Cancer Research Panel was designed to detect expression imbalance in fusion driver genes using only 10 ng of FFPE RNA. The panel is a single pool of primers targeting fusion transcripts included in the ALK, RET, ROS1, and NTRK genes.

Ion AmpliSeq RNA Cancer Panel

The Ion AmpliSeq™ RNA Cancer Panel was developed as an RNA analysis complement to the Ion AmpliSeq™ Cancer Hotspot Panel v2. The panel is a single pool of primers targeting 50 oncogenes and tumor suppressor genes including the KRAS, BRAF, and EGFR genes.

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Ion AmpliSeq RNA Apoptosis Panel

The Ion AmpliSeq™ RNA Apoptosis Panel is an analysis tool designed against 267 TaqMan Assay–verified genes involved in the cellular apoptosis pathway. Included are genes associated with death receptor–mediated apoptosis, c-myc-mediated apoptosis, and p53-mediated apoptosis.

Targeted RNA sequencing workflow for the Ion PGM or Ion S5 systems
1 Select/design
targeted panel
Enrich RNA Library

Select from our ready-to-use gene panels or submit your own set of genes online and we will design and create a customized Ion AmpliSeq panel just for you.

2 Construct
Construct library for RNA-Seq

• Perform reverse transcription and amplify targets in a single reaction
• Digest primer sequences and ligate Ion Torrent specific adapter sequences

3 Prepare
Prepare Template for RNA-Seq

• Load prepared library, prepackaged reagents, and up to 2 Ion chips onto Ion Chef instrument
• Start run for automated template preparation and chip loading

Learn more about Ion Chef System ›

4 Run
Run Sequence for RNA Sequencing

• Initialize Ion PGM or Ion S5 instrument
• Load sequencing-ready chip from Ion Chef System and start sequencing run

Learn more about Ion PGM & Ion S5 system ›

5 Analyze data and validate
Analyze data for RNA Sequencing

• Perform automated analysis using Torrent Suite Software with the Coverage Analysis plug-in to generate transcript count data
• More extensive analysis can be performed using Partek Flow Software

Targeted RNA sequencing informatics solutions

Torrent Suite Software coordinates all the experiment planning and data processing steps necessary to complete your targeted sequencing workflow. New product templates enable users to plan targeted runs simply and quickly. In just a few hours, raw data are captured on the Ion PGM or Ion S5 sequencer and transferred to the Torrent Server for base calling. Digital counting is then completed using the AmpliSeqRNA plug-in, which reports normalized transcript counts in spreadsheet file formats, enabling users to import data into other software packages for differential expression analysis. A more comprehensive analysis can be performed using Partek Flow Software.

Excellent correlation with TaqMan Assays

Universal human reference (UHR) RNA and human brain reference (HBR) RNA control samples were used to compare fold change for each of the 50 genes included in the Ion AmpliSeq RNA Cancer Panel against the fold change calculations of the same genes with TaqMan Gene Expression Assays. A resulting correlation of 0.989 was observed. These results show the accuracy of the gene expression data obtained with the Ion AmpliSeq RNA workflow. Find TaqMan Assays corresponding to the same gene regions interrogated by Ion AmpliSeq RNA amplicons in the Ion Community.


* Based on recommendations of multiplexing 4-6 libraries generated using the Ion AmpliSeq™ RNA Apoptosis Panel on one 318™ chip.
**The content provided herein may relate to products that have not been officially released and is subject to change without notice.