peak in the physiological dinucleosome region only between 90 and 110 kb. At 120 kb the Amplitude at the Period value of 337 bp diminishes slightly, and the peaks at 380 and 415 bp increase relative to those in the 100 kb window. Thus, the regular, in-phase oscillations in period-10 VWG are present in large-scale chromatin structures. The Fourier Amplitude reaches maximum strength at 70 kb or greater and remains strong until about 100 kb for most of the loci studied. In two cases the signal reached maximum intensity at 90 kb and remained strong until 200 kb. In one case a strong signal was detectable at 40 kb and it persisted until 500 kb. In addition to the ten tested predictions, we have included several other entries in Table 1. The ﬁt parameters and signal characteristics are listed for the previously studied mada gene locus (17), a locus predicting a 191 bp repeat, and a locus predicting a 210 bp repeat. The 191 bp prediction was con- ﬁrmed, but the very high 210 bp prediction was not. Addi- tionally, we included two probes from loci that did not contain a signal; these are representative of most of the genome. These probes detected the bulk repeat, as predicted. We also included the analysis of a ladder that resulted from a probe that did not hybridize speciﬁcally, but detected a very large number of loci (detecting a continuum of restriction fragment sizes, instead of the single intended fragment). This probe provided a way of measuring the bulk repeat by Southern blotting, as used in these experiments, instead of by traditional gel stain- ing. The ﬁt parameters obtained by either method were very similar. Statistical significance of the results confirming our computational predictions The percentage of mouse liver chromatin possessing nucleo- some repeats shorter than 180 bp can not be large because, if it were, the effect of having such repeats present in appreciable amounts should be readily detectable. This point is illustrated in Figure 3 which shows a simulated 195 bp ladder (195) next to a simulated 180 bp ladder (180) and the superposition of these two ladders (merged). It can be seen that the oligomer bands greater than the 5mers become out of phase with each other. This ‘vernier effect’ causes the superimposed ‘ladder’ to lose resolution at about the 6mer where the bands of each individual ladder become maximally out of phase with each Figure 3. Simulated nucleosome ladders having two different repeats illustrat- ing the vernier effect. A ladder having 195 bp spacing is shown adjacent to a ladder having 180 bp spacing. At least 10 bands can be resolved for each. The superposition of these two ladders (merged) leads to a loss of resolution after the 5mer due to a vernier effect. Nucleic Acids Research, 2006, Vol. 34, No. 7 1979 other. For short repeat values that are less than 180 bp, the loss of resolution is even more severe (data not shown). Because the bulk chromatin ladder from mouse liver does not lose resolution until beyond the 10mer (17), it can be immediately concluded that the percentage of mouse liver chromatin having repeats shorter than about 180 bp is considerably less than 50% (the percentage in the simulated superimposed ladders of Figure 3). The statistical signiﬁcance of our predictions (Table 1) can be assessed if the percentage of the chromatin in mouse liver nuclei that possesses short nucleosome repeats is known. To estimate this percentage experimentally, we made use of the vernier effect illustrated above. We made mixtures of DNA from MNase-digested mouse liver chromatin (195 ± 5 bp bulk repeat) and M.incana petal chromatin (bulk 183 ± 5 bp repeat) with Matthiola petal: mouse liver weight ratios of: 50:50, 40:60, 30:70, 20:80 and 10:90. We ran these ladder mixtures on an agarose gel along with pure mouse liver chromatin and pure Matthiola petal chromatin nucleosome ladders (Figure 4A). It can be seen that the pure mouse liver (ML) and pure Matthiola petal (MP) nucleosome ladders each extend to the 10mer or beyond, and the ladders begin to go out of phase with each other after the 5mers. As expected, the 50:50-mixture gives a vernier effect similar to the simulation shown in Figure 3. It can also be seen that the vernier effect is detectable at least down to the 20:80 mixture. In Figure 4B the upper regions of the ladders for the 20:80 mixture and for mouse liver chromatin are shown expanded and next to each other. Lane scans are shown in Figure 4C which conﬁrm the visual impression that the peaks for the nucleosome oli- gomer bands greater than the 5mer are better resolved for the pure mouse liver chromatin than for the 20:80 mixture. Thus, a conservative estimate for the maximum percentage of short repeat chromatin in mouse liver nuclei is about 20%, and therefore the probability of selecting a hybridization probe by chance that detects a short repeat is 0.2. We think that this number is representative of both the repetitive and non- repetitive portions of the genome because 20% of the non- speciﬁc probes, which hybridized to repetitive DNA, detected repeats that were shorter than the bulk value. The probability of selecting 10 out of 10 probes that detect short repeats (as in Table 1) by chance alone is then about (0.2)10 ¼ 0.0000001, or one in ten million. Hence, it is highly unlikely that 10 out of 10 probes detecting short nucleosome repeats were selected simply by chance, rather than by following our computational predictions, and the results reported in Table 1 are therefore statistically signiﬁcant. DISCUSSION Our results demonstrate that loci exist in mouse liver chro- matin with nucleosome repeats that are well below the bulk chromatin value. Moreover, our results strongly suggest that these loci can be identiﬁed from a computational analysis of the DNA sequence. This is the ﬁrst time that non-repetitive DNA sequence has been used to predict an aspect of large-scale chromatin structure. Experimentally, we have estimated that less than 20% of the chromatin in mouse liver nuclei possesses nucleosome repeats that are <180 bp, values at least 15 bp shorter than the bulk chromatin repeat. This estimate is roughly consistent with our computational