RNA amplification techniques enable researchers to profile small clinical samples, laser capture microdissected
(LCM) samples, sorted cells and even single cell samples using microarrays. Amplification kits are also available
for formalin-fixed parrafin-embedded (FFPE) tissue samples and other partially degraded RNA samples. All RNA samples
processed at the PMGC are assessed for quality by the Agilent Bioanalyzer. Once the quality of the RNA sample has
been assessed, the most appropriate RNA amplification method for the platform can be determined.
The PMGC uses various RNA amplification techniques (listed below) for labelling picogram to nanogram quantities
of total RNA.
NuGEN Amplification and Labelling Systems
several amplification systems based on their proprietary Single Primer Isothermal Amplification (Ribo-SPIA™) technology.
The Ovation® RNA amplification systems (Ovation Amp, WT-Ovation Amp, WT-Ovation Pico, WT-Ovation FFPE V2) linearly
amplify all mRNA species, including low abundance transcripts. NuGEN also offers the Applause™ System for preparing
targets for analysis on Affymetrix GeneChips, and offers target preparation modules specific for the Affymetrix, Agilent,
and Illumina platforms.
Minimum amount of total RNA required: 500 pg (using WT-Ovation® Pico kit V2); 50 ng (using Applause™)
Sigma Amplification (TransPlex WTA)
TransPlex Whole Transcriptome Amplification (WTA) Kit
uses a blend of quasi-random primers to ensure accurate
transcriptome coverage and rapid amplification.
Minimum amount of total RNA required: 5 ng
Ambion RNA Amplification (MessageAmp)
The MessageAmp™ II aRNA Amplification Kit
based on antisense RNA (aRNA) amplification first described by
Van Gelder and Eberwine
, includes the patented MEGAscript™ technology for high-yield in vitro
Minimum amount of total RNA required: 100 ng (single round of amplification); 0.1 to 100 ng (two rounds)
Iscove Global RT-PCR Amplification
A global RT-PCR amplification method, described by Dr. Norman Iscove
(Nature Biotechnology 2002, 20:940
uses a rapid and highly optimised global RT-PCR procedure to exponentially amplify RNA yet preserve abundance relationships.
Minimum amount of total RNA required: 20 pg
- Can partially degraded samples be amplified?
In some cases, partially degraded total RNA samples can also be amplified. Generally, more total RNA is required for partially degraded samples.
- What is tyramide signal amplification (TSA)?
Tyramide signal amplification (TSA) is a patented signal amplification technology that amplifies the fluorescent signal. The TSA method can increase the detection sensitivity up to 100-fold.