Chemical structure and biological activity 



Caplin, 1951), and artichoke tuber tissue can be grown in almost exactly the 

 same fashion as carrot tissue. 



Use is now being made of another technique which was devised primarily 

 to permit relatively large amounts of cultured tissue to be obtained for the 

 purposes of biochemical examination. Here the tube, normally containing 

 one or at most two or three individual carrot explants, is replaced by a flask 

 {see Figure l{a)). Around the periphery of the flask, nipples are blown in such 

 a way that the tissue explants distribute themselves in these projections and 

 again are alternately exposed to air and bathed in liquid. 100 explants from 

 an individual carrot root can be successfully grown in a culture of this sort 

 containing 250 ml of nutrient fluid, and in this fashion weights up to 10 g 

 of tissue can be grown in about 28 days. The standard deviation of such a 

 population of explants is very small indeed (zb^-lO per cent of the mean). 

 However, an unexpected sequel to this technique is of interest. Explants 

 taken from the mature carrot root grown by this 'flask technique' grow in 

 much the same fashion as individual explants in a normal culture tube, that 

 is, they produce more or less ellipsoidal masses of a relatively undifferentiated 

 parenchymatous type of tissue. However, when concentrated in the flask the 

 supernatant becomes somewhat turbid, owing to the development of free 

 floating cells which grow in the medium. Such floating cells can be used to 

 make liquid inocula into other flasks. This technique has been in continuous 

 use now for some two years and it has become a common experience that a 

 large number of small tissue cultures can be grown from these liquid inocula 

 which contain only free floating cells, either singly or in very small groups. 

 Also, quite contrary to expectations it was found that cultures grown from 

 these free floating cells are much more prone to organize and form roots than 

 they are when grown in the normal medium from explants newly isolated 

 from the carrot root. Reference is made to this technique here, however, 

 merely to emphasize that it is now quite clear that the tissue culture technique 

 can be extended downward so that inocula can be made, not only from small 

 tissue explants from the whole organ, but also from a stock of free growing 

 cells. 



GROWTH REqUIREMENTS OF DIFFERENT TISSUES 



Carrot and artichoke tissue grow adequately when a normal basal medium 

 is supplemented by the addition of whole coco-nut milk. The condition, 

 however, is quite different in the case of the potato tuber. Depending 

 somewhat on the individual tuber and the strain from which it was derived, 

 virtually no external growth occurs when minute pieces of potato tissue are 

 exposed to the conditions under which carrot tissue grows apace. To produce 

 an actively growing tissue culture of potato tuber, a synergistic mixture is 

 necessary. This consists of 2 parts: (a) the coco-nut milk complex, and 

 (b) 2:4-D or an equivalent substance present in minute amount in the solu- 

 tion. For 2:4-D itself, the most effective concentration is of the order of 

 6 p. p.m. in a medium containing about 10 per cent by volume of whole 

 coco-nut milk (Steward and Caplin, 1951). In this curious twofold require- 

 ment for substances that induce proliferative growth in the cells of the potato 

 tuber, it is now clear that both substances are continuously necessary. This 

 can be shown by the experiments illustrated in Figure 2, in which the tissue 



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