[Properties of replicating DNA in the regenerating rat liver isolated during phenol fractionation].

Using phenol fractionation in the absence of detergents three DNA fractions differing by the incorporation of radioactive thymidine after pulse label are obtained from regenerating rat liver cells. Two fractions extracted under variety of conditions represent the main bulk of cell DNA (85--90%). DNA non-extractable under conditions used (DNA III) incorporates labelled thymidine 10--15 times faster than the first two DNA fractions. DNA III purified from the interphase layer by pronase, sodium dodecylsulfate and phenol sediments at 26S and has a hyperchromic effect about 40% after alkaline denaturation. Alkaline sucrose density gradient centrifugation of pulse-labelled DNA III revealed that nascent DNA consisted of heterogeneous fragments similar in size to the replication fragments in bacterial cells (9--10S). It was shown by CsCl equilibrium centrifugation that buoyant density of heat denatured DNA III labelled for 5 min with [3H]thymidine is heavier than the bulk of DNA prelabelled for 2 h with [14C]thymidine. After hydrolysis with RNase or alkali, buoyant densities of both DNAs became the same. These results support the idea of initiating role of RNA in the synthesis of discontinuous replicating fragments in regenerating rat liver cells. Specific radioactivity of RNA associated with replication fragments which are labelled for 5 min with [14C] orotic acid is 20 times more than of the same RNA labelled for 30 min. These data demonstrate high metabolic activity of initiating RNA.