Analysis of highly purified satellite DNA containing chromatin from the mouse

Abstract

A purification scheme for satellite DNA containing chromatin from mouse liver has been developed. It is based on the highly condensed state of the satellite chromatin and also takes advantage of its resistance to digestion by certain restriction nucleases. Nuclei are first treated with micrococcal nuclease and the satellite chromatin enriched 3-5 fold by extraction of the digested nuclei under appropriate conditions. Further purification is achieved by digestion of the chromatin with a restriction nuclease that leaves satellite DNA largely intact but degrades non-satellite DNA extensively. In subsequent sucrose gradient centrifugation the rapidly sedimenting chromatin contains more than 70% satellite DNA. This material has the same histone composition as bulk chromatin. No significant differences were detected in an analysis of minor histone variants. Nonhistone proteins are present only in very low amounts in the satellite chromatin fraction, notably the HMG proteins are strongly depleted.