I. REPLICATION OF DNA IN CELL-FREE SYSTEMS 29 



analyzed by Schachman et al. (1958) and in eveiy parameter tested 

 they resembled native calf thymus DNA. In Table XII the viscosities, 

 sedimentation constants, and molecular weights for synthesized DNA 

 and calf thymus DNA (used as primer) are compared, and it can be 

 seen that the two materials are closely related. jMolecular weights of 

 several preparations range from 4 to 6 million and the viscosity and 

 sedimentation data indicate that the deoxynucleotide units are organized 

 in the macromolecule to form relatively stiff rods with effective volmnes 

 greater than would be expected from single polynucleotide chains with 

 freedom of rotation at each internucleotide link. If the product is heated 

 at 100°C for 15 minutes (Table XII) there is a sharp drop in viscosity 

 with a relatively small decrease in sedimentation. When the product is 

 digested with DNase there is a 30% increase in extinction, or hyper- 

 chromic effect (Fig. 8). These latter two observations support the concept 

 of an organized rigid arrangement of deoxyribonucleotides in the super- 

 structure of the native molecule. 



F. PRIMER-PRODUCT RELATIONSHIPS 



As mentioned in the introduction, the tenn "replication" is taken to 

 mean the synthesis of a DNA which is in every way indistinguishable 

 from the primer. In this sense it would be better to use the word "tem- 

 plate" rather than primer to describe the small amount of DNA required 

 for the synthesis of more DNA since this implies a copying mechanism 

 rather than initiating mechanism as the function of the added DNA 

 although, the two functions go hand in hand, except in some instances 

 mentioned later. "Primer" will be used throughout this discussion for 

 convenience, but it should be borne in mind that the primer used in 

 DNA synthesis is unique in biochemistiy in that it serves both as initia- 

 tor and template. 



So far we have seen that in respect to gross chemical structure and 

 physical profile the synthetic DNA closely resembles the primer. We 

 would like to be able to determine the exact sequence of deoxyribo- 

 nucleotides in the product and compare this to the primer molecule; but, 

 at present, techniques are not available for doing this even for relatively 

 small polynucleotides. Valuable information has been obtained toward 

 elucidating the fine structure of the synthetic DNA by determining its 

 base composition and dinucleotide frequencies. 



1. Base Composition 



Two general methods have been used to determine the base composi- 

 tion of the newly formed DNA. In the first procedure four reaction 

 mixtures identical except for the radioactive deoxyribonucleoside tri- 



