248 PLANT GROWTH SUBSTANCES 



auxin to exposed surface is supposed to account for the much more 

 widespread and more pronounced enlargement obtained with relatively 

 large explants. 



Where present, this initial phase is commonly followed by the for- 

 mation of an irregular cambium immediately below the surface, often 

 involving cells which have undergone some enlargement.This cambium 

 cuts off cells to the outside which are rapidly suberized. There is 

 usually little or no new internal tissue formed. The cells of the interior 

 of the fragment become woody, chlorophyll is formed in the outer- 

 most layers if the explants are exposed to light, and the fragment, 

 although it may enlarge to as much as six or eight times its original 

 volume, finally settles into a static condition in which it may remain 

 alive and green but not growing for several months. If small fragments 

 have been used enlargement is generally quite regular, the original 

 outlines of the explant being recognizable after many months. Large 

 fragments may give rise to localized, tubercular growths along with 

 nongrowing or necrotic areas. If such a fragment is divided into two 

 or four pieces and the parts are replanted on fresh substratum, the cut 

 surfaces will undergo a similar series of changes but will again come to 

 rest. The presence or absence of externally supplied nutritive substances, 

 sugars, salts, and the like, apparently plays no role in this phase of de- 

 velopment, which is entirely dependent, except for the moisture 

 required, on residual suppUes within the explant. It is sometimes pos- 

 sible to continue this subdivision through five or six passages, but it 

 is not possible to establish continuously growing strains. This de- 

 velopmental pattern has been observed repeatedly since the days of 

 Haberlandt and Rechinger. It is quite consistent and has led more than 

 one early investigator to make premature claims to having established 

 tissue cultures, before it was realized that a few such passages are not 

 sufficient to establish the ability of such cultures to grow continuously. 

 Neither Gautheret (1934 — Salix) nor myself (1939 — tomato and beet) 

 has been free from this fault. 



In 1939, however, Gautheret and Nobecourt independently showed 

 that, if a growth substance such as indoleacetic acid or naphthalene- 

 acetic acid is added at a suitable concentration to a complete nutrient 

 either at the time of the initial excision or as a substratum for the first 

 subcultures (that is to say after completion of the initial residual 

 growth) when growth becomes dependent on external materials the 



