(A-T) develops which has the physical size and properties of natural DNA 

 and in which the adenine and thymine are in a perfectly alternating sequence. 

 When this rare form of DNA-Hke polymer is used as a primer, new A-T 

 polymer synthesis starts immediately and even though all four triphosphates 

 be present, no trace of guanine or cytosine can be detected in the product. The 

 conclusion from these several experiments thus seems inescapable that the base 

 composition is replicated in the enzymatic synthesis and that hydrogen-bonding 

 of adenine to thymine and guanine to cytosine is the guiding mechanism. 



Enzymatic replication of nucleotide sequences. 



The fourth line of evidence which I would like to cite is drawn from current 

 studies of base sequences in DNA and their replication. As I have suggested 

 already, we believe that DNA is the genetic code; the four kinds of nucleotides 

 make up a four-letter alphabet and their sequence spells out the message. At 

 present we do not know the sequence; what Sanger has done for peptide 

 sequence in protein remains to be done for nucleic acids. The problem is more 

 difficult, but not insoluble. 



Our present attempts at determining the nucleotide sequences (33) will be 

 described in detail elsewhere and I will only summarize them here. DNA is 

 enzymatically synthesized using P^^ ^s label in one of the deoxynucleoside 

 triphosphates; the other three substrates are unlabeled. This radioactive 

 phosphate, attached to the 5-carbon of the deoxyribose, now becomes the 

 bridge between that substrate molecule and the nucleotide at the growing 

 end of the chain with which it has reacted (Fig. 9). At the end of the synthetic 

 reaction (after some 10^® diester bonds have been formed), the DNA is isolated 

 and digested enzymatically to yield the 3' deoxynucleotides quantitatively. 

 It is apparent (Fig. 9) that the P atom formerly attached to the 5-carbon of 

 the deoxynucleoside triphosphate substrate is now attached to the 3-carbon 

 of the nucleotide with which it reacted during the course of synthesis of the 

 DNA chains. The P^^ content of each of the 3' deoxynucleotides, isolated by 

 paper electrophoresis, is a measure of the relative frequency with which a 

 particular substrate reacted with each of the four available nucleotides in 

 the course of synthesis of the DNA chains. This procedure carried out four 

 times, using in turn a different labeled substrate, yields the relative frequencies 

 of all the sixteen possible kinds of dinucleotide (nearest neighbor) sequences. 



Such studies have to date been carried out using DNA primer samples from 

 six different natural sources. The conclusions are: 



i) All 16 possible dinucleotide sequences are found in each case. 



S-61 



