I. REPLICATION OF DNA IN CELL-FREE SYSTEMS 43 



with no appreciable lag and continues linearly. When dGTP or both 

 dGTP and dCTP are omitted from the incubation, there is a 4-hour lag 

 followed by a period of rapid synthesis. If no triphosphates are added to 

 the incubation mixture, or just those indicated in the last curve in the 

 figure, no synthesis occurs. The lag period can be reduced somewhat by 

 raising the enzyme concentration; but it can be eliminated completely 

 by priming the reaction with synthetic d-AT polymer (Fig. 14). Only 

 d-AT polymer itself is an effective primer in the reaction, and other 

 DNA preparations have no effect in reducing the lag period. 



When the "limited" reaction (Section IV,C) is investigated, using 

 d-AT as the primer, small but significant quantities of deoxyadenylate 

 or deoxythymidylate are added to the ends of the polymer. It is of 

 interest that neither dCIVIF nor dGMP can add to the ends of the d-AT 

 polymer, suggesting that even in the limited reaction specific base-pairing 

 is required. This requirement for specific base-pairing favors the "repair" 

 hypothesis suggested as one of the possible explanations of the "limited" 

 reaction (Section IV,C). 



Analysis of the d-AT polymer shows it to be a rodlike organized 

 structure with a molecular weight ranging between 10'^' and 10". It is 

 composed exclusively of equal proportions of deoxyadenylate and deoxy- 

 thymidylate residues, and an interesting feature is that these residues 



TABLE XX 

 Hydrolysis of d-AT Copolymer to S'-Deoxyribonucleotides"'^ 



° From Schachman et al. (1960). 



''d-AT copolymer was hydrolyzed to 3'-deoxyribonucleotides by micrococcal DNase 

 and splenic phosphodiesterase until more than 96% of the P'- was released in the form 

 of deoxj'ribonucleoside monophosphates. 



are arranged in a strictly alternating sequence. This is demonstrated in 

 Table XX where the results of a nearest neighbor analysis show that 

 each deoxythymidylate residue is linked by a phosphodiester bridge to 

 a deoxyadenylate residue and vice versa. This regularly alternating 

 sequence has been used to explain the obser^'ation that the d-AT co- 



