The biosynthesis of protein amino acids in plant tissue culture I. Isotope competition experiments using glucose-U-C14 and the protein amino acids.

Abstract

Most investigators have used excised tissue, cellfree preparations, or whole plants for studies of metabolism in angiosperms. Each of these systems represents a different state of the cells or tissues. Another state of plant cells, that of exponential growth, has been obtained by tissue culture techniques (2, 5). Studies of the metabolism of cells growing exponentially would provide information which is complementary to that already available for plants. Such studies would allow direct comparison of metabolism of higher plant cells with that of microorganisms also growing exponentially. When specific information is not available, it is generally assumed that the metabolism in plant cells is the same as that in micro-organisms. This assumption may be tested using exponentially growing plant cells. For use in studies of metabolism a culture system in which plant cells are growing exponentially should have several features. The media used should have a known composition. The tissue should be grown in suspension as very small clumps or single cells. The use of media of known composition makes it possible to manipulate the nutrients at will and to avoid the presence of unrecognized compounds which occur in media supplemented with natural extracts. The growth of tissues as suspensions of single cells or small clumps of cells prevents the formation of the diffusion gradients which occur in larger pieces of tissue. Rapid agitation of the suspension keeps the cells in equilibrium with the medium and the medium in equilibrium with its gaseous atmosphere. Such desirable features can now be found in at least 2 plant tissue cultures. These are cultures established from Paul's Scarlet Rose by Dr. W. Tulecke (Tulecke, private communication) and from Nicotiana tabaccumn var. Xanthi by Dr. P. Filner (22). Methods established for growth of microbial cultures have been successfully adapted to the culture of plant cells. For example, plant tissues have been maintained on solid media as callus. Single plant cells have been plated in agar to yield visible colonies (8). Also, plant tissues have been maintained as suspensions of cells and clumps in liquid media of known or unknown composition (2) in shake flasks or large carboys (29). The close parallels in methods of growth suggest that methods developed to study the nietabolism of microbial cells in culture might also be readily adapted to the study of plant cell metabolism. Several studies (10, 18, 34) of plant metabolism have employed the isotope competition method (25). This paper is to report the application of the isotope competition method to the study of amino acid biosynthesis in suspensions of plant cells growing exponentially in media of known composition.