Figure 5.

(A) Single- and (B) double-stranded gels of a polynucleosome sample (larger than 20 nucleosomes) that was digested with DNase I, either in the presence of MnCl₂ (A, lanes b–e; B, lanes c–f), in the presence of MgCl₂ (A, lanes f–k; B, lanes g–l) or in the presence of MgCl₂ after mild trypsin digestion (A, lanes f and g, B, lanes g and h). Samples were digested with 2, 4, 8 or 16 U DNase I/mg DNA at 37°C. In (A) lanes f and g and (B) lanes g and h samples were digested with 2 µg trypsin/mg DNA at room temperature for 2 min (D, lane 2). The reactions were terminated with 40 µg trypsin inhibitor/mg DNA and 1 mM PMSF final, after which digestion was performed with 2 or 4 U DNase I on ice for 15 min. (C) Double-stranded gel of a DNase I-digested mixture of tri- and tetranucleosomes, which were not digested with trypsin and thus have H1/H5 histones present. They were digested with 0, 1, 2, 4, 6 or 8 U DNase I/mg DNA. (D) SDS–PAGE of histones from the chromatin samples digested with 2 mg trypsin/mg DNA for 0, 2, 4, 8, 12 or 16 min (lanes 1–6, respectively). GH1/5, globular parts of H1/H5 histones.