Secondary structure
Secondary structure of small RNA molecules is largely determined by strong, local interactions such as hydrogen bonds and base stacking. Summing the free energy for such interactions should provide an approximation for the stability of a given structure. To predict the folding free energy of a given secondary structure, an empirical nearest-neighbor model is used. In the nearest neighbor model the free energy change for each motif depends on the sequence of the motif and of its closest base-pairs. The model and parameters of minimal energy for Watson–Crick pairs, GU pairs and loop regions were derived from empirical calorimetric experiments, the most up-to-date parameters were published in 2004, although most software packages use the prior set assembled in 1999.