Distribution of RNA polymerase binding sites in fractionated chromatin

Calf-thymus chromatin was fractionated by ion-exchange chromatography on ECTHAM-cellulose and sucrose gradient sedimentation. [ECTHAM-cellulose is a cationic adsorbent prepared by coupling tris(hydroxymethyl)aminomethane to cellulose with epichlorohydrin.] The capacity of these fractionated chromatins to support RNA synthesis by DNA-dependent RNA polymerase of Escherichia coli was examined, using procedures that permit measurement of binding site frequency. Unfractionated calf thymus chromatin has 5-10% as many binding sites as protein-free DNA. By combination of the two fractionation methods, chromatin samples were obtained containing as few as 2% and as many as 47% of the number of binding sites found on protein-free DNA.