DotKnot-PW has been designed as a dedicated pseudoknot prediction tool and should be applied to RNA sequences where pseudoknotted interactions are suspected in the structures. Prediction accuracy will inevitably decrease for sequences, which are longer than say 400 nt for any single sequence structure prediction method (Reeder et al., 2006) . To achieve reliable results, short sequences should be folded using DotKnot and predictions should be compared with results from other methods from the literature. To gain confidence in predictions, subsequent comparative prediction using DotKnot-PW and other comparative methods is highly recommended. Ideally, experimental verification of computationally predicted pseudoknots should be sought. Fig. 3 . Pairwise prediction results for the S15 mRNA pseudoknot (RFAM family RF00114) with the top two combined free energy and similarity scores. The reference structure is shown at the top and folds into two conformations in dynamic equilibrium: a H-type pseudoknot or a series of hairpins.