smith: directed mutation 425 



cursors; (c) incorporation of these unnatural analogues into DNA 

 during synthesis of the latter; and (d) occurrence of permanent but 

 nonlethal errors (mutations) in the base sequence during subsequent 

 replication of the modified DNA. It is conceivable that if nitrogen 

 mustards alkylate purines (Zamenhof) and ethylene imines pyrimi- 

 dines (Szybalski), a basis may be provided for mutagenic specificity 

 between these two classes of alkylating agents. 



An ultimate step in demonstrating the mechanism of mutagenic 

 specificity would be to show that a hereditary change induced by 

 altering the nucleic acid has, in turn, produced a change in amino 

 acid sequence of associated protein. Tsugita and Fraenkel-Conrat 

 (67) have reported that the protein of a differential host mutant 

 isolated after nitrous acid treatment and reconstitution of tobacco 

 mosaic virus (TMV) RNA differs from that of the parent strain in 

 that three amino acid residues are replaced by three others (proline, 

 aspartic acid, and threonine by leucine, alanine, and serine). 



Mutagenic Specificity and 

 Directed Mutation in Higher Organisms 



Experiments on microorganisms, fungi, bacteria, and bacterio- 

 phage, have provided the unequivocal evidence for mutagenic specific- 

 ity. These analyses have, for the most part, made use of techniques 

 developed to explore the genetic fine structure of complex loci (14). 

 It has been shown that specificity is characteristic of alleles at numer- 

 ous sites in the gene locus rather than of the locus in general. This 

 intra-locus or inter-allelic specificity has been demonstrated in Neuro- 

 spora and bacteria only by the back-mutation test; in phage by both 

 forward ("hot spot" spectra) and reverse mutations. Auerbach and 

 Westergaard (5) suggest that forward mutations involve damage or 

 malfunction by diverse mechanisms and are therefore less strictly 

 specific; whereas, repair of a damaged allele would require a specific 

 mechanism. Two mutagens which show an allele-specific effect in a 

 back-mutation test may not show any striking specificity in a broad 

 spectrum forward-mutation experiment. 



A question of primary interest is. to what extent can the results 

 with microorganisms be related or applied to higher organisms 

 There are differences in the organization of the heritable material 

 from phage to the chromosomes of higher forms in strand composi- 



