Complementary techniques: RNA amplification for gene profiling analysis

The study of clinical samples is often limited by the amount of material available. DNA and RNA can be amplified from small specimens and, therefore, used for high-throughput analyses. While precise estimates of the level of DNA concentration in a given specimen is rarely studied (with the exception of relatively crude analyses of gene amplification or loss in cancer specimens), it is critical to know the proportional expression of various RNA transcripts since this proportion governs cell function by modulating the expression of various proteins. In addition, accurate estimates of relative RNA expression in biological conditions portray the reaction of cells to environmental stimuli shedding light on the characteristics of the microenvironment associated with particular physiologic or pathologic conditions. For this reason, the development of technologies for high fidelity messenger RNA amplification have been focused of extreme interest in the past decade with specific aim not only of increasing the abundance of RNA available to study but to accurately maintain the proportionality of expression of various RNA species among each other within a given specimen. This chapter will discuss various approaches to proportional RNA amplification focusing on amplification of the whole transcriptome (all transcripts in a given samples) rather than individual genes. These methods are suitable for high-throughput transcriptional profiling studies.