IV. CELLULAR CONTROL OF DNA BIOSYNTHESIS 157 



DNA synthesis and the beginning of mitosis as well as the duration of 

 the period of DNA synthesis. (For a detailed review of the design and 

 interpretation of such experiments see Taylor, 1960a.) 



Figure 2 is an example of such an experiment in which exponential 

 multiplying L cells (tissue culture strain originally derived from the 

 mouse) were labeled with thymidine-H^ and the peak grain count at 

 metaphase measured at intervals thereafter. As may be seen, no counts 

 were recorded for 3^ hours after the onset of labeling ; this is the period 

 (Go) between the cessation of DNA synthesis and metaphase. The num- 

 ber of grains/metaphase nucleus then rises for 6-7 hours after which 

 no increase in labeling is observed; this, then, is the period of DNA 

 synthesis, or S, during which incorporation of thymidine can occur. The 

 number of grains will be low for those nuclei which were leaving S 

 when thymidine-H'^ was introduced. The grain number per nucleus will 

 rise until a maximum value is reached which represents nuclei which 

 have passed through their entire S period in the presence of the radio- 

 active isotope. The duration of S is thus given by the length of time 

 during which the grain count/nucleus rises. vSince the generation time of 

 these cells was 20 hours and mitosis occupies a period of 1/3 hour, Gi 

 may be estimated as between 9 and 11 hours. Note the sharply defined 

 appearance of labeled mitoses and of the saturation of the grain count, 

 indicating the similar behavior of different individuals of the random 

 population. 



The cycle of DNA synthesis may also be reconstructed from micro- 

 spectrophotometric measurements of the DNA content of individual 

 cells sampled at random (Swift, 1950; Walker and Yates, 1952; Alfert, 

 1955; Bloch and Godman, 1955; Vendrely and Vendrely, 1956; Richards 

 et al, 1956; McDonald, 1958; Walker and Mitchison, 1958; Kimball 

 and Borka, 1959; Smith et al, 1959; Painter et al, 1960; McLeish and 

 Sunderland, 1961; Utsume, 1961). Such reconstructions have been sub- 

 stantiated by determination of the DNA content of individual cells at 

 known intervals following mitosis and by measurements of synchronous 

 cultures. 



Figure 3 represents an example of such an experiment using embry- 

 onic mouse heart cells grown in tissue culture. Two observations are 

 noteworthy: (a) the similarity between the DNA cycle obtained by this 

 method and that illustrated in Fig. 2, and (6) the precise control of the 

 quantity of DNA within the cells, the number of cells containing 

 amounts of DNA less than 22 or greater than 44 units being negligible. 



Finally, experiments on synchronous cell populations have measured 

 the total amount of DNA in the population and assumed that this would 

 reflect the course of synthesis by a single cell (Ogur et al, 1953; Hotch- 



