46 Essays in Biochemistry 



urably; there were 20% more survivors in an aliquot irradiated in its 

 starvation medium than in one suspended in fresh deficient medium. 

 Thus, under the conditions of these experiments, the culture of 10 8 cells 

 per milliliter being washed before the start of the starvation, nucleic 

 acid fragments in the medium contribute very little to inaptitude. In 

 the experiments in which we first demonstrated protection by starva- 

 tion media, 10 9 cells per milliliter were starved for 4 hours and, after 

 elimination of the organisms by sterile filtration, cells in logarithmic 

 growth phase were added to the filtrate to yield a final concentration 

 of 10 8 cells per milliliter. The protection by screening was thus more 

 pronounced. 



However, it should be emphasized that the protection by the starva- 

 tion medium is not due merely to screening, for if such media are 

 added to bacteria after the irradiation they still suppress the induction : 

 the number of colony-forming survivors is increased five- to eightfold, 

 and there is a decrease in the number of infectious centers. The 

 starvation medium can be added, in a ratio of 1 part to 10, as late 

 as 20 minutes after the irradiation, and a significant reversal of induc- 

 tion is still measurable. The starvation medium has no effect on the 

 non-lysogenic E. coli B/r after its irradiation. 



As yet we know no more about this induction-reversing agent than 

 that it is dialyzable and is labile to ultraviolet irradiation. 



Since so much of nucleic acid fragments are excreted during starva- 

 tion the question naturally arises whether inaptitude may not be the 

 result of intracellular shading of radiation-sensitive loci by these frag- 

 ments as they flow towards the periphery of the bacterial cell. To 

 explore such a possibility, the nucleic acid content of the methionine 

 deficient auxotroph was studied after two types of starvation, glucose 

 and methionine. 



For these studies 100-ml. aliquots of either glucose- or methionine- 

 starved organisms were centrifuged at intervals in a Sorvall angle 

 centrifuge at 5000 r.p.m. for 20 minutes. A cell count in the superna- 

 tant fluid revealed the presence of 1%, or less, of the original bacteria. 

 This finding, together with the finding that there was no change in the 

 number of viable, colony-forming bacteria during the course of the 

 starvations, insured the uniformity of sampling at various intervals. 

 The bacterial pellet containing 2 to 3 X 10 10 colony-forming cells was 

 washed once with 0.9% saline, centrifuged, and subjected to analysis 

 for RNA, DNA, and dilute-acid-soluble fragments by the method of 

 Ogur. 



