RELEASE OF GENETIC TRANSFORMING AGENT FROM PNEUMOCOCCAL CULTURES DURING GROWTH AND DISINTEGRATION

Abstract

Genetic transformations of pneumococcus mediated by streptomycin-induced lysates were studied to gain some insight into the nature of freshly released transforming principle, and the influence of the physiologic state of the donor population on the transformation process. It was found that streptomycin could make the DNA of sensitive cells available for transformation of other cells. Living cultures of pneumococcus growing exponentially in ordinary media were also found to discharge significant quantities of genetically active DNA. Such cultures, not treated with any drug, showed no evidence of concomitant cell disintegration or death. Both single markers and small linkage groups could be transferred in transformations mediated by drug-induced lysates and by filtrates of living cultures. The quantity of DNA liberated is small (less than 0.1 µg per ml), but these transformations are at least as efficient as transformations mediated by purified DNA, when compared on the basis of total DNA available. Up to 1 per cent of the cells in an average recipient culture can be transformed by a small quantity of culture fluid. Both in drug-treated and in untreated cultures the amount of transforming activity increased and then decreased during growth of the culture. Although the source of transforming DNA in growing cultures could not be established, the decline in the transforming activity of aging drug-treated or untreated cultures was attributed to the presence of deoxyribonuclease. The release of this nuclease by pneumococcal cultures midway in exponential growth is sufficient to result in a mild degradation of the low concentration of freshly released transforming agent present. Maximal release of active transforming agent by a living culture coincided in time with the development of maximal receptivity to exogenous DNA by that culture. As a result, recombinants could be recovered from appropriately genetically marked strains growing in each other's presence. In view of these results, it seems possible that DNA-mediated transformations might provide, or might have provided, a mechanism of genetic recombination in nature for some bacterial species in which sexual mechanisms may not be available.