Abstract
A new two-step fractionation procedure has been developed which rapidly but quantitatively and qualitatively separates HeLa cell nuclear proteins from nuclear DNA and RNA. Purified HeLa cell nuclei, chromatin (prepared by two different procedures), or isolated heterogeneous ribonucleoprotein particles were first dissociated with sodium dodecyl sulfate detergent, and nuclear proteins and nuclear nucleic acid were partitioned after the addition of sodium chloride and chloroform:isoamyl alcohol. When isolated nuclei of HeLa cells (pulse labeled with [ 3H]leucine) were subjected to this fractionation procedure, over 95% of the radiolabeled nuclear protein was found in the combined interphase and organic phase of the extraction mixture. In parallel experiments, over 90% of the 3H-uridine labeled nuclear RNA and 95% of the [ 3H]thymidine labeled nuclear DNA were consistently partitioned into the aqueous phase of the centrifuged suspensions. Acid soluble nuclear proteins (mostly histone), acid insoluble nuclear proteins (nonhistone), or total nuclear proteins were prepared utilizing this procedure and were analyzed by sodium dodecyl sulfate polyacryalmide gel electrophoresis.
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