Studies on Transformations of Hemophilus influenzae

Abstract

A B S T R A C T A procedure has been developed for obtainingHem0philus influenzae of such competence that 1 to 10 per cent transform to any of several genetic factors by utilizing a period of aerobic growth followed by a non-aerobic period. Differences in levels of competence were not due to differences in genetic background. Competence was due to at least one factor intrinsic to the cell or site on the cell and was not transferable to non-competent cells. Competence was affected by salt concentration, pH, and temperature. Washing competent cells reduces their ability to transform, but not their capacity to bind DNA reversibly. The irreversible step could be restored with little or no accompanying growth. These facts suggest that reversible and irreversible binding represent separate biochemical steps. DNA initiates a reaction in cells leading to a loss of competence. In the absence of DNA the cells remain competent for at least an hour. Competence correlates quantitatively with predictability of multiple transformations. The observed and calculated values of multiple transformations are in closer agreement, the higher the frequency of transformation for single markers. The correction needed to bring the two figures into agreement is a measure of the fraction of non-competent ceils.

Transformability of Haemophilus influenzae

“…injluenxue strain Rd. This observation seems to disagree with a report by Goodgal & Herriott (1961) who state that the anaerobically held bacteria (in our study : sensitized) are competent. We believe that in their experiments bacteria may have become competent during the relatively long exposure to DNA.…”

“…This fraction is close to one in highly competent cultures, i.e. cultures showing transformation frequencies of one per cent or more (Stuy, 1 9 6 1~; Goodgal & Herriott, 1961). As a consequence, we feel confident to state that the actual number of competent bacteria is roughly 100 times the number of transformants observed in cases where their frequencies are lower than one per cent.…”

“…1958) obtained competent cultures of Haemophilus in$umzae through interruption of the aerobic growth of a culture by a period of more or less anaerobic incubation. This was recently described in detail (Goodgal & Herriott, 1961). Stuy (1960), however, pointed out that holding aerobically grown bacteria (in vigorously shaken suspensions) under conditions excluding oxygen nearly completely, was merely a 'pretreatment'.…”

Autoradiographic Estimation of Competence and the Relationship between Competence and Transformability in Cultures of Bacillus subtilis

“…As was to be expected the calculated size of the competent fraction is found to be independent of DNA concentration or the time of exposure to DNA, which follows from the expression (A x B)/C from which the competent fraction is calculated (Goodgal & Herriott, 1961). If the frequencies of the two types of single transformants increase by a factorp and q respectively, the size of the competent fraction will be ( P A x qB)/pqC.…”

Competence in Bacillus subtilis Transformation System

“…Many bacterial species are able to take up deoxyribonucleic acid (DNA) when in a state of 'competence' and can then be transformed (Alexander & Redman, 1953; Klein & Klein, 1953;Balassa, 1954;Bracco, Krauss, Roe & MacLeod, 1957;Corey & Stan, 1957;Hotchkiss, 1957;Spizizen, 1958 ;Goodgal & Herriott, 1961 ;Catlin & Cunningham, 1964). The DNA derived from bacteriophage (Romig, 1962;Green, 1964;Okubo, Strauss & Stodolsky, 1964;Reilly & Spizizen, 1965) or virus (Abel & Trautner, 1964;Bayreuther & Romig, 1964) can similarly be incorporated in competent bacteria where it can replicate.…”