Prepartation of soluble chromatin and specific chromatin fractions with restriction nucleases

Abstract

By digestion of rat liver nuclei with EndoR HaeIII, EndoR EcoRI, and EndoR Bam and subsequent lysis of the nuclei approx. 90%, 40%, and 45%, respectively, of the chromatin were solubilized. The plateau values of solubilization are in agreement with a model in which the chromatin strands are crosslinked and/or attached to a supporting structure. The distribution of DNA lengths in the soluble and insoluble chromatin fractions were determined. According to digestion experiments with restriction nucleases rat liver DNA contains highly repetitive sequences, some of which are arranged in tandem repeats of 95 and 380 nucleotide pairs, respectively. With EndoR EcoRI chromatin containing the repetitive RNA was preferentially solubilized and, by subsequent sucrose gradient centrifugation, purified to about 90%. The useful properties of chromatin prepared by the specific action of restriction nucleases are discussed.