General Appearance, Growth Pattern, and Anatomical Structure of Crown-Gall Tissue of Nicotiana tabacum L. grown in vitro on culture media containing Glucose or soluble Starch as a Carbon Source

Substitution of glucose as a carbon source for soluble starch can lead to a striking alteration in the general appearance of tissue cultures. This phenomenon was observed in tobacco crown-gall tissue and in a strain derived from the cambial zone of twigs of Rubus fruticosus L. (Karstens and De Meester-Manger Cats, 1960). In the present paper details are given on the general appearance, growth pattern, and anatomical structure of tobacco crown-gall tissue cultures. In Table I differences of shape, colour, consistency, and manner of growth in relation to external conditions have been summarized. A method is described by which it is possible to study the growth pattern of tissue cultures: initial fragments or young tissue cultures are dusted with powdered charcoal under sterile conditions. After growth has taken place, the localization of the charcoal particles can be studied macroscopically, or microscopically in sections prepared according to routine microtechnical methods. Besides the tobacco cultures on glucose and starch media under varied external conditions, a number of other tissue cultures were tested. As a result, several growth types could be distinguished. It is a matter of some interest that the tissue cultures of tobacco crown-gall on glucose and starch media possess totally different growth habits. In the glucose cultures the initial fragment remains intact and exhibits a very special differentiation while in the starch cultures the initial fragment “dissolves” into the growing culture. In glucose cultures new growth is strictly localized, i.e. in a meristematic zone formed at the outer surface of the initial fragment, the side in contact with the culture medium being excepted. In starch cultures, however, growth takes place by the activity of scattered meristematic loci present in the initial fragment and afterwards in the whole tissue culture. The initial fragments in glucose cultures, as mentioned before, exhibit a very special differentiation. In the initial fragment a great number of cup-shaped meristematic zones are initiated and give rise to complexes of heavily lignified cells. In the newly-formed tissue lignification takes place to a much lesser extent and in such a way that lignified elements are sparsely present in otherwise parenchymatic tissue. In starch cultures cup-shaped meristems occur very rarely. They can only be found in light cultures. In addition, few lignified elements are formed by those meristems. On the whole, starch cultures grown either in the light or in the dark have a parenchymatic character with few and very scattered lignified elements. Finally, the lignification pattern of the lignified elements proved to be different in glucose and starch cultures. The anatomical features agree very well with the figures on dry weight/fresh weight ratios given in a previous paper (Karstens and De Meester-Manger Cats, 1960) and summarized in Table II.