VII. SYNTHESIS OF RNA AND RIBOSOMES 337 



appreciable deviations from randomness occur in DNA (Josse et al., 

 1961). Alkaline hydrolysis should then release nucleotides which show 

 an apparent composition equal to the composition of newly formed RNA 

 if the RNA is similar in its distribution of neighbors to ribosomal RNA. 

 If, however, the RNA is similar to DNA and contains a non-random 

 distribution, the apparent composition will not be an accurate measure 

 of the newly formed material. 



At early times after the addition of P^- the radioactivity of the free 

 nucleotide pool is much greater than that of the nucleic acid. Some 

 of this radioactivity is likely to contaminate the nucleic acid. It is 

 therefore necessary to use separation techniques capable of separating 

 the 2',3'-nucleotides released by alkaline hydrolysis from contaminating 

 5' nucleotides or serious errors may result. Also, to minimize errors from 

 losses or conversions (by deamination) during hydrolysis it is desirable 

 to measure the radioactivity by isotope dilution (Fig. 28). 



B. TIME COURSE OF COMPOSITION 



Differences in the compositions of newly formed material have been 

 observed in several situations. Volkin and Astrachan found the RNA 

 formed after infection by T2 was intermediate in composition between 

 the RNA of the bacteria and the DNA of the phage (Volkin and 

 Astrachan, 1956). The formation of RNA approaching the composition 

 of the DNA was observed in several bacteria (of quite different DNA 

 composition) following a transition from broth to sjmthetic medium 

 (Hayashi and Spiegelman, 1961). 



The same effect also appears in the absence of intentional transients 

 in the growth conditions. Yeas and Vincent observed the apparent com- 

 position of the RNA formed by yeast during 5-20 minute periods of 

 exposure to P'*'- (Yeas and Vincent, 1960). Although the data scatter 

 widely, the apparent composition seems to be intermediate between the 

 RNA of the cell and its DNA. Similar results were obtained in a variety 

 of bacteria (Astrachan and Fisher, 1961). 



An extensive series of measurements has been carried out by ]\Iidgley 

 using several different organisms (jMidgley, 1962) (Table VI). The 

 accuracy of these measurements, carried out by isotope dilution, is 

 sufficient to allow an estimate of the time constant of the transition. 

 Correcting for the delays introduced by the nucleotide pool, the time 

 constant is less than 3 minutes, corresponding roughly to that of the 

 eosome stage of ribosome synthesis. Also the composition of the newly 

 formed material (extrapolated to zero time) is not like that of the DNA 

 but is midway between the DNA and the RNA of the cell. 



