x, matching nt; N, wobble of A, C, G, or T. even better. By restricting the wobbles to 3 (e.g. a D or a B) or 2 (e.g. a Y or a T) nt, the probability of a match becomes even greater. Thus, in a highly degenerated probe with at least one continuous region of perfectly matching nt, a large part of the pool will extend this region and contribute to nucleation, zipping and hybridization. The behavior of the highly degenerated probes is encouraging and in accordance with the NucZip model, which predicts that the high likelihood of several matching stretches of 5 nt or longer will result in significant hybridization. One of the aims of this study was to investigate the binding capacity of dInosine in 3M TMAC. 5-NitroIndole was chosen as a comparative universal base. The results shown in Figure 3C demonstrate that 5-NitroIndole in the 5-NitroInd_18 probe had a much greater destabilizing effect than dInosine in the Ino_18 probe, without the same capacity to rescue hybridization with a target containing many sm. Furthermore, the 5-NitroInd_18 probe was more affected than the Ino_18 probe when the hybridization temperature was raised from 45 to 55 C. It is concluded that, under 3 M TMAC buffer conditions, dInosine is a better choice than 5-NitroIndole when designing a variation-tolerant probe with as little degeneration as possible.