274 M. K. Bach and J. Fellig 



applied at relatively high concentrations (3). Under such circimi- 

 stances, the application of plant growth regulators must disrupt the 

 control mechanisms which are normally existent and cause the pro- 

 liferative growth of the tissue. It seemed, therefore, that such a sys- 

 tem might be suitable for an investigation of the immediate bio- 

 chemical changes which take place upon addition of a plant growth 

 regulator, in the hope of finding the chemical causes for the initiation 

 of proliferative growth. It is recognized, of course, that these changes 

 cannot be equated to the physiological mode of action of the plant 

 growth regulators. However, the interest here is primarily in the 

 chemical or enzymatic changes which are involved in the disruption 

 of normal growth and the initiation of proliferation, and the use of 

 ])Iant growth regulators to bring about these changes is somewhat 

 incidental. Indeed, the initiation of proliferation by a variety of 

 chemicals (not necessarily plant growth regulators) has been recog- 

 nized for a long time (6). 



The main difficulty in such an investigation is the critical differ- 

 entiation between the causes and the effects of proliferation. Most 

 of the reported effects of 2,4-dichlorophenoxyacetic acid (2,4-D) on 

 biochemical systems (4, 15, 16, 24) have involved lengthy in vivo pre- 

 treatments. It is not at all clear whether these effects are the conse- 

 quences rather than the primary causes of the morphological and 

 physiological changes which are induced by this compound. These 

 difficulties can be circumvented by approaching the biochemical 

 studies from a kinetic point of view — investigating the rate at which 

 the biochemical changes take place in the very early stages of pro- 

 liferation, with a view to establishing a precursor-product relationship 

 between the changes themselves. However, before such a sttidy can 

 be undertaken, it is essential that the time interval involved in the 

 initiation of proliferation be clearly defined. To this end it must be 

 established how long and in what form the inducing agent persists 

 ill the tissue, what concentrations of inducing agent are required, 

 and what conditions must be met for the initiation of proliferation 

 to take place. 



This paper will describe some of our results in trying to define 

 these variables. Since beans are particularly susceptible to callus forma- 

 tion (20, 21), they were used in this study. 



MATERIALS AND METHODS 



Phaseohis vulgaris, 'Giant Stringless Potl Bean,' was grown in the 

 greenhouse in potting soil between 18 and 33° C. until 3 or 4 trifoliate 

 leaves had fully developed. The plants were harvested just before 

 use, the leaves removed, and the stems immersed in water until used. 



