Bacteria: Recombination {V) 



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When these two strains were mixed and 

 then plated on medium lacking methionine 

 and threonine, prototrophic colonies ap- 

 peared in such large numbers that they 

 could be explained entirely, or almost entire- 

 ly, as being the result of genetic recombina- 

 tion, and not of mutation. Prototrophs also 

 were obtained, if a liquid culture of the M+T' 

 strain was centrifuged (to remove most of the 

 bacteria), the supernate heated for 20 to 30 

 minutes (to kill any remaining bacteria), and 

 the supernate added to the M T+ strain. This 

 demonstrates that no living M+T^ donor cells 

 are required in order to furnish the M+ factor 

 for the estabhshment of prototrophy. So, 

 this is clearly not a case of genetic recombina- 

 tion involving conjugation. Moreover, the 

 filtrate retained its full M+ capacity after 

 treatment with DNAase. Accordingly, this is 

 not a case of genetic recombination via trans- 

 formation. Since the M+ factor could pass 

 through filters that held back bacteria but not 

 viruses, it can be called a filterable agent (FA). 

 It was also found that the M+T- strain 

 harbored a phage. This virus, P22, is said to 

 be nonvirident or temperate, for only occasion- 

 ally does it become virulent, at which time it 

 replicates itself and then lyses or bursts the 

 host cell to liberate as many as several hun- 

 dred progeny phage. Accordingly, temperate 

 phage does not cause conspicuous lysis. Since 

 the M+T~ strain of Salmonella carries P22 as 

 a temperate virus and is potentially capable of 

 having its cells lysed, it is said to be a lyso- 

 genic strain. Lysogeny also confers on the 

 bacterium immunity to infection by identical 

 or homologous phage. The MT^ strain 

 happens normally to lack P22, that is, it is a 

 nonlysogenic or sensitive strain. When a 

 sensitive strain is infected with temperate 

 phage, a certain fraction of the cells lyse and 

 liberate phage, but another fraction survives, 

 becomes lysogenic, and gives rise to lysogenic 

 progeny. Lysogenic bacteria can be lysed 

 artificially and tested for phage. None is 

 detected. Apparently, in cells that become 



lysogenic, the phage is converted to a new 

 form, called prophage, which reproduces at 

 the same rate as the host. When a lysogenic 

 cell is to be lysed by viral action, prophage 

 normally first rapidly replicates a number of 

 times to produce infective phage which is 

 liberated at the time of lysis. 



A natural question to ask now is: What is 

 the relationship between the filterable agent 

 M+ and the phage P22? The following 

 facts were determined experimentally. Both 

 FA A/+ and phage P22 (1) were unafi'ected 

 by RNAase and DNAase, (2) showed the 

 same inactivation pattern with temperature 

 changes, (3) had the same susceptibility to an 

 antiserum that blocks the attachment of phage 

 to the bacterium, (4) attached to susceptible 

 cells simultaneously, (5) had the same size 

 and mass as determined by filtration and 

 sedimentation tests, (6) appeared in the 

 medium at the same time and in a constant 

 ratio, and (7) retained this constant ratio 

 even though various purification and concen- 

 tration procedures were applied. It is evident 

 from these results, therefore, that FA M+ is 

 phage-bound. Since the genetic material of 

 Salmonella is known to be composed of DNA, 

 so is the genetic factor M+. Moreover, since 

 fact (1) precludes the attachment of the M+ 

 genetic factor to the outer surface of the 

 phage particle, the M+ gene must be located 

 in the interior of the virus. 



Genetic transduction is the term we shall use 

 to define the process of genetic recombination 

 which is made possible when homologous 

 DNA, contained within a virus particle, is 

 sent into a recipient cell. 



Is there any limitation to the genetic ma- 

 terial which can be transduced by P22? P22 

 can be grown on bacteria which are geneti- 

 cally marked M+T+X+Y~Z-, and the crop of 

 phage produced, following this infection, can 

 be harvested. Part of the harvested phage is 

 then tested on suitable indicator strains (M~, 

 T-, X-, Y-, Z-) one at a time. The results 

 show transduction of M+, of T+, and of A'+j 



