Targeted RNA Sequencing by Ion Torrent Next-Generation Sequencing
Ion AmpliSeq™ gene panels
Targeted, quantitative gene expression analysis.
Read the application note describing the high quality data generated by Ion AmpliSeq™ RNA technology
See corresponding TaqMan® Assays to Ion AmpliSeq™ RNA panels
Empower your data analysis with push-button analysis providing number of reads per targeted gene sequence.
Targeted RNA workflow for the Ion PGM™ System
Design your own Ion AmpliSeq™ RNA Custom Panel
Creating and ordering custom panels is easy with the 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 simply enter your specific genomic content online and we will design and create customized Ion AmpliSeq™ Panel just for you. Leveraging more than a decade of expertise powering the TaqMan® Custom Assay design pipeline, AmpliSeq™ Designer produces optimized primer designs in just hours. You now have full flexibility to analyze hundreds of genes of your choice, such as those implicated in a particular disease state or representing specific biochemical pathways. Ion AmpliSeq™ RNA custom designs target selection of 12 to 300 human amplicons in a single reaction, leveraging the experience from a community of >1000 DNA-based AmpliSeq™ registered users with >5,000 panels submitted. High-quality amplicons are designed to account for GC content, and filtered for repeats and SNPs. Ion AmpliSeq™ RNA amplicons are designed to target exons that represent the maximum number of transcripts for single genes that have multiple transcript isoforms documented in standard databases such as RefSeq.
In addition to custom panels, there are two Ion AmpliSeq™ Ready-to-use Panels that were designed and validated by Life Technologies:
Ion AmpliSeq™ RNA Cancer Panel
The Ion AmpliSeq™ RNA Cancer Panel was developed as a RNA 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 KRAS, BRAF, and EGFR genes.
Ion AmpliSeq™ RNA Apoptosis Panel
The Ion AmpliSeq™ Apoptosis Panel is a screening tool designed against 267, TaqMan® validated, genes involved in the cellular apoptosis pathway. Included are genes associated with death receptor– mediated apoptosis, c-myc, and p53-mediated apoptosis.
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. Raw data are processed on the Ion PGM™ Sequencer and transferred to the PGM™ Torrent Server for base calling and variant calling. This entire process takes just a few hours, and further analysis can be performed using the Coverage Analysis plug-in which reports amplicon counts in spreadsheet file formats which allow users to import data into other software packages for differential expression analysis. A more comprehensive analysis can be performed using Partek® Flow software.
Overview of Targeted RNA Sequencing
Researchers employ targeted sequencing as a rapid and cost-effective alternative to whole genome sequencing and focus on selected sets of genes or genomic regions. The ability to efficiently and evenly amplify just the genomic regions of interest and to keep the RNA input amount low are critical factors, especially in human disease research applications. Expression profiling is of fundamental interest to a wide range of biological research applications including gene regulation analysis, correlating gene expression with phenotypic information, and cellular pathway analyses. Targeted RNA sequencing, using Ion AmpliSeq™ technology, 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 formalin-fixed, paraffin-embedded (FFPE) samples. Further, targeted RNA sequencing can be used to detect fusion transcripts from chromosomal rearrangements that are critical initiating steps of tumorigenesis. Generate the data equivalent of up to 1600 individual TaqMan® qPCR assays in a single Ion PGM™ run. This technology, coupled with the Ion PGM™ System, Torrent Suite™ Software and the Coverage Analysis plug-in, enables researchers to go from precious FFPE samples to quantitative gene expression profiling in a single day.
* 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.
For Research Use Only. Not for use in diagnostic procedures.
The Ion Total RNA-Seq Kit v2 provides a fast, simple workflow with as little as 100 ng of total RNA input, while maintaining strand orientation and minimizing bias and error. The Ribominus™ Eukaryote Kit v2 provides more than 90% rRNA depletion increasing the sequencing power of the Ion Proton™ System. The Ambion® ERCC RNA Spike-In Mix comprises a set of unlabeled, polyadenylated transcripts designed to be added to an RNA analysis experiment after sample isolation in order to measure dynamic range, the lower limit of detection, and fold-change response.