Abstract
A general method for isolation and fractionation of chromatin into its four major components, DNA, RNA, histories, and nonhistone proteins, is described. The procedure avoids the use of strongly acidic or alkaline conditions, or the use of ionic detergents or phenol. As few as 14 × 106 cells can be used. The procedure is reasonably rapid and has been used successfully with a number of tissue culture cell lines. The chromatin components are dissociated in a 3 M NaCl – 5 M urea solution containing 2-mercaptoethanol and EDTA. The DNA and high molecular weight RNA are collected by high-speed centrifugation and DNA is separated from the RNA by means of Cs2SO4 equilibrium density centrifugation. The histones, nonhistone proteins, and low molecular weight RNA's are fractionated using DEAE-cellulose column chromatography and polyacrylamide gel electrophoresis. A small amount (< 1%) of protein is present in the DNA and RNA fractions. At least 11 low molecular weight RNA subfractions can be detected by means of polyacrylamide gel electrophoresis.
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