VII. SYNTHESIS OF RNA AND RIBOSOMES 315 



phases of incorporation into nucleic acids might be misinterpreted as 

 indicating an exchanging component of the nucleic acid. 



V. Kinetics of Incorporation into RXA of Ribosomes 



Kinetic studies of ribosome synthesis depend upon methods of sepa- 

 rating various classes of macromolecules. The techniques which have 

 proved to be most useful are chromatography and sedimentation anal- 

 ysis. The latter technique can be used both for the analysis of the 

 ribonucleoprotein particles and of the RXA extracted from them. 



In the following paragraphs details have been drawn exclusively from 

 the work carried out in our laboratoiy (Roberts et al., 1958; McCarthy 

 and Aronson, 1961; McCarthy and Britten. 1962: Britten et a/.. 1962 1. 

 The reasons for this are twofold. On the one hand, we are more familiar 

 with our own work, and on the other, different experiments can be 

 related more surely because they were carried out under similar 

 conditions. Experiments in other laboratories also contribute to our 

 understanding. 



An RXA fraction of high specific radioactivity has been separated 

 by electrophoresis (X'omura et al., 1960). Otaka et al. have observed 

 precursor product relationships among fractions separated on DEAE 

 (Otaka et al.. 1962 1. Gros et al. have studied an RXA component of 

 high specific radioactivity which is dissociated from ribosomes when the 

 magnesium concentration is reduced and which has an average sedi- 

 mentation coeflBcient of about 12S (Gros et al.. 19611. 



A. DEAE chrgmatogr-aphy 



Figure 13 shows the elution patterns obtained from cells given 

 progressively longer exposures to C^^-uracil. After breaking the cells, the 

 walls and membranes were removed by a brief period of centrifugation. 

 Ribosomes and their precursors were sedimented by prolonged centrifu- 

 gation i40K 240' I leaving the S-RX'A in the supernatant fluid. This 

 procedure is desirable because S-RXA elutes at 0.5 M X'aCl contami- 

 nating ribosome precursors. DXA. which also elutes in this region, was 

 removed by DX'ase. 



The patterns show clearly that the C^^-uracil first appears in ma- 

 terial eluting at 0.6 -V XaCl. After the first few minutes this component 

 reaches a saturation level and C^*-uracil builds up in material eluting 

 at 0.5 -1/ XaCl. As time goes on the second region approaches saturation 

 and C^*-uracil appears in the bulk of the ribosomes which elute at 0.4 M 

 X'aCl. The column seems to have separated two sequential precursors 

 from the final product. This qualitative impression is confirmed by the 



