ROBERTS, BRITTEN, AND BOLTON 



91 



a nonradioactive medium after exposure to the tracer. In this treatment any 

 intermediates which have a rapid turnover should lose their radioactivity. The 

 resulting "persistent pattern" was entirely similar to the "steady-state pattern," 

 and no protein components could be identified as intermediates. 



Finally, cells were exposed to the tracer for short periods. After a 4-minute 

 exposure the resulting "pulse pattern" was similar to the "steady-state pattern" 

 except that the radioactivity of the nucleoprotein peak was only one-half of that 

 expected from the "steady-state" pattern. A similar result was obtained with 

 cells exposed for 4 minutes to a mixture of C 14 -labeled amino acids. 



Similar experiments carried out with P 32 Oi give much more striking results. 

 Figure 6 shows the macromolecular region of the elution patterns obtained with 

 cells exposed to the tracer for increasing periods of time. The radioactivity ap- 

 pears first in a quite distinct fraction of the elution pattern, passing through at 

 a later time to the other regions. In the steady-state and persistent patterns the 

 phosphorus radioactivity was proportional to the optical density (at 260 mu). 

 Thus the DEAE column is capable of resolving the nucleic acid and nucleo- 

 protein into fractions that seem to be precursors and products. Similar kinetic 

 differences were also observed by Creaser, who used ECTEOLA columns [9] 

 to analyze alcohol-extracted nucleic acid [14]. 



The analysis of these data runs into a number of complications. The leading 

 peak is composed solely of RNP, but the secondary peak is an unresolved mix- 

 ture of RNP, RNA, and DNA. Furthermore, the pool of low-molecular-weight 

 precursors to RNA is large and may or may not be in equilibrium with the 

 smaller pool of DNA precursors [14]. 



A rough analysis can be made on the basis of several simplifying assump- 

 tions. Assume first that the low-molecular-weight precursors of the macromole- 



DE AE COLUMN ANALYSIS OF E.COLI AT EARLY TIMES AFTER ADDITION OF P 32 



I 1/2 mm 



6min 



24 min 



30 



Fraction number 



Fig. 6. Elution patterns of cell extracts after growing cells were exposed to P 32 4 for 

 times indicated. Only a small region of the elution pattern is shown. 



