The concept of NucZip, with both local and distal cooperativity contributions, is an attempt to predict the hybridization behavior of most 70-mer probe-target combinations under the given conditions ( Figures 6A and 8) . The NucZip algorithm is now under revision to include probes containing 1, 2, 3 or 4 nt wobbles, as well as taking into account the results with the universal base 5-nitroindole. The algorithms, schematically described in Figure 8 , (i) were based on our experimental data (described above) and (ii) included highly matching nucleation sites extending beyond the neighboring nt. Thus, unlike the NN theory, NucZip takes the effects of matches at longer distances into account. The unique property of 3 M TMAC to provide a roughly equal contribution to hybridization by A:T and G:C pairs justifies simple computational approaches. The well known additivity of binding contributions per nt inherent in ÁG calculations according to the NN theory (22, 45) indicates that hybridization can be treated in a relatively simplistic way. Our concepts were based on the finding that the longer the sequence of uninterrupted matching nt, the more stable is the hybridization. Our calculations were thus focused more on binding than on destabilization, i.e. the use of positive rather than negative contributions. One of the weaknesses of the NN theory is that it adds all contributions, positive or negative, to a grand sum. The negative contributions are subtracted for the whole molecule, instead of in the local context where they belong. In our approach, the binding is first assessed locally, mimicking nucleation, and then extended cooperatively to the whole molecule, mimicking the zipping process. It is binding that keeps the hybrid together, and it is thus logical to focus on binding.