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RNA interference, mediated by small interfering RNAs (siRNAs), is a powerful tool for investigation of gene functions and it is increasingly being used as a therapeutic agent. However, not all siRNAs are equally potent - although simple rules for the selection of good siRNAs were proposed early on, siRNAs are still plagued with widely fluctuating efficiency. Recently, new design tools that incorporate both the structural features of the targeted RNAs and the sequence features of the siRNAs have substantially improved the efficacy of siRNAs. In this chapter, we present the algorithms behind these accessibility-aided tools and show how to design efficient siRNAs with their help.

RNA interference (RNAi) has become an important tool to study and utilize gene silencing by introducing short interfering RNA (siRNA). In order to predict the most efficient siRNAs, a new software tool, RNA Workbench (RNAWB), has been designed and is freely available (after registration) on http://www.rnaworkbench.com. In addition to the standard selection rules, RNAWB includes the possibility of statistical analyses of the applied selection rules (criteria). The role of RNA secondary structures in the RNA interference process as well as the application of sequence rules are discussed to show the applicability of the software.

Background: Reliable prediction of RNA-RNA binding energies is crucial, e.g. for the understanding on RNAi, microRNA-mRNA binding and antisense interactions. The thermodynamics of such RNA-RNA interactions can be understood as the sum of two energy contributions: (1) the energy necessary to 'open' the binding site and (2) the energy gained from hybridization., Methods: We present an extension of the standard partition function approach to RNA secondary structures that computes the probabilities Pu[i, j] that a sequence interval [i, j] is unpaired., Results: Comparison with experimental data shows that Pu[i, j] can be applied as a significant determinant of local target site accessibility for RNA interference (RNAi). Furthermore, these quantities can be used to rigorously determine binding free energies of short oligomers to large mRNA targets. The resource consumption is comparable with a single partition function computation for the large target molecule. We can show that RNAi efficiency correlates well with the binding energies of siRNAs to their respective mRNA target., Availability: RNAup will be distributed as part of the Vienna RNA Package, www.tbi.univie.ac.at/~ivo/RNA/