The incorporation of [3H]cytidine into ribonucleic acid of liver nuclei of young and old rats

Abstract

The ribonucleic acid of rat-liver nuclei labeled with [³H]cytidine and harvested from liver homogenates using a phenol-NaCl mixture may be fractionated by successive extractions at temperatures between 20 and 75°, using a phenol-NaCl extraction medium. Successive extractions at any one temperature produce extracts whose ribonucleic acid content decreases markedly. The analysis of the specific activity of extracted nuclear ribonucleic acid indicates two fractions: (1) a relatively low specific-activity fraction obtained at 25, 40 and 50°; and (2) a relatively high specific-activity fraction obtained at 65 and 75°, temperatures at which measurable amounts of deoxyribonucleic acid are also extracted.

It is demonstrated that InCl₃ at appropriate concentration and ionic strength, effectively precipitates all soluble ribonucleic acid as well as all deoxyribonucleic acid as the indic nucleates. The indic nucleates are solubilized in 0.38 N KOH which results in the precipitation of In (OH)₃ in an exchange reaction that is complete in about 2 h at room temperature. In this way the ribonucleic acid may be hydrolysed to constituent nucleotides and the unhydrolysed deoxyribonucleic acid may be recovered by perchloric acid precipitation.

The results of extraction experiments with nuclei harvested from liver of young and old adult rats indicate: (1) the specific activity of ribonucleic acid extracted from “old” liver nuclei is consistently higher than that extracted from “young” nuclei; (2) the amount of ribonucleic acid extracted from “young” liver nuclei is greater than that extracted from “old” liver nuclei; and (3) the amounts of deoxyribonucleic acid extracted from “young” and “old” liver nuclei are approximately equal.