158 M. J. Thornley, J. Sinai and J. Yudkin 



method of Boivin were not able to transform sensitive cells, 

 although various modifications of the conditions of prepara- 

 tion were made. 



The method of McCarty and Avery consists of lysis with 

 deoxycholate in the presence of citrate, removal of protein 

 with chloroform and amyl alcohol and precipitation of the 

 transforming principle by absolute alcohol. Four rough 

 strains, derived from the original smooth strain, were tested 

 for competence to undergo transformation. In one of these, 

 there was a slight increase in the number of proflavine- 

 resistant cells. We thought that this might be due to the 

 existence of small numbers of a substrain consisting of com- 

 petent cells. We therefore devised a method of "double 

 replica plating", by which we might isolate them. A master 

 plate of one of the rough strains was replicated on plates con- 

 taining the transforming principle. After incubation, this in 

 turn was replicated on plates containing proflavine. Colonies 

 on this plate were traced back to corresponding areas of the 

 master plate which had been kept at 5°. From ten of these, 

 subcultures were made and transformation attempted. In 

 one of them, transformation was achieved as indicated by a 

 200-fold increase in the number of resistant cells in presence 

 of DNA from resistant cells, but no increase in presence of 

 DNA from sensitive cells. Two further experiments with the 

 same rough strain, and one with a different rough strain, gave 

 similar results. 



The third method, that of Myers and Spizizen, was used by 

 these authors to produce a highly polymerized DNA, although 

 they did not carry out transformation experiments with it. 

 The method consists of lysing with sodium dodecyl sulphate 

 (duponol), removal of protein with sodium acetate and pre- 

 cipitation of DNA with acidified alcohol. Transformation 

 was tested on the original smooth strain of Esch. coli. The 

 DNA preparation alone did not cause transformation, the 

 protein precipitate caused transformation in about 0-3 per 

 cent of the cells, and both preparations together caused trans- 

 formation in ten times as many cells. No transformation 



