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RNAi: A How To for New Users


RNA interference, the biological mechanism by which double-stranded RNA (dsRNA) induces gene silencing by targeting complementary mRNA for degradation, is revolutionizing the way researchers study gene function. For the first time, scientists can quickly and easily reduce the expression of a particular gene in mammalian cell systems, often by 90% or greater, to analyze the effect that gene has on cellular function. The ease of the technique, as well as the wide availability of high quality kits and reagents for performing RNAi, have contributed to its rapid adoption by the research community. This article provides an overview of RNAi, the requirements for a typical RNAi experiment, and the Ambion products that simplify each step.


The RNAi Mechanism
In non-mammalian systems, introducing or expressing long double-stranded RNA (dsRNA) triggers the RNAi pathway. The cytoplasmic nuclease Dicer first cleaves the long dsRNA into 2123 bp small interfering RNAs (siRNAs), that then unwind and assemble into RNA-induced silencing complexes (RISCs) (Figure 1). The antisense siRNA strand then guides the RISC to complementary RNA molecules, and the RISC cleaves the mRNA, leading to specific gene silencing. However, since most mammalian cells mount a potent antiviral response upon introduction of dsRNA longer than 30 bp, researchers transfect cells with 2123 bp siRNAs to induce RNAi in these systems without eliciting the antiviral response.

Figure 1. Overview of RNA Interference.


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