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Nuclear and cytoplasmic RNA from sheep thyroid slices in vitro labeled with [3H]uridine or [32P]phosphate from 5 min up to 9 h, have been obtained by different phenol extraction procedures. Early-labeled RNA was only extracted by phenol containing 0.1 or 1% (w/v) sodium dodecylsulfate (SDS) at 0° or 60°, whereas stable nuclear or polysomal RNA was easily extracted by phenol at 0°. Time course of incorporation of label into subcellular particles has shown the early labeling of nuclear RNA and the transfer of radioactivity into the cytoplasm. Rapidly-labeled RNA extracted from nuclei by phenol-1% (w/v) SDS and analyzed by sedimentation was shown to sediment in three discrete classes 50–30 S, 22 S and 13 S, different from ribosomal RNA. After 9 h of incubation the 22 S and 13 S labeled RNAs were still present in detectable amounts in the nuclei, indicating a somewhat low turnover rate for these components. The kinetics of incorporation of label into polysomes was studied in thyroid slices incubated from 15 rain up to 5 h with [3H]uridine or [32P]phosphate. RNA was dissociated from polysomes by dodecylsulfate treatment and centrifuged on sucrose gradient. As for nuclei, the rapidly-labeled RNAs sediment in discrete size classes, 22 S and 13 S. After a long incubation time (5 h) the label was predominant in 28 S and 18 S ribosomal RNA but 28 S RNA was not completely labeled. Up to 30 min labeling a high molecular weight, early-labeled RNA was present in purified polysomes. Its radioactivity declines after 60 rain labeling. A part of the rapidly labeled polysomal RNA can be reversibly separated from rRNP by the action of EDTA (1.25 to 5 μmoles EDTA/mg RNP). It sediments in a broad zone (12–30 S) of high specific radioactivity. Increasing Mg++ concentration results in the reassociation of rapidly-labeled RNA to ribosomal sub-particles. Buoyant densities of the polysomal RNP particles obtained by EDTA treatment of 32P-labeled polysomes and stabilized by means of formaldehyde fixation, were determined by banding in CsCl gradients. The bulk of the stable material banded at characteristic densities (d = 1.55 and 1.48), but most of the rapidly-labeled RNA banded at d = 1.38. The specific radioactivity of this material was the same as that of the 12–30 S RNA isolated by sucrose gradient centrifugation from the same EDTA­treated polysomes. Preliminary double labeling experiments and computation of the RNA to protein ratio calculated from the RNA and protein density values indicate this material to be early-labeled nucleoprotein particles with a high protein content. [ABSTRACT FROM AUTHOR]