GROWTH INDUCTION IN CULTURES OF HAPLOPAPPUS GRACILIS. I. THE BEHAVIOR OF THE CULTURED CELLS

BLAKELY, L. M., and F. C. STEWARD. (Cornell U., Ithaca, New York.) Growth induction in cultures of Haplopappus gracilis. I. The behavior of the cultured cells. Amer. Jour. Bot. 48(5): 351358. Illus. 1961.-Because of the unusual cytology of Haplopappus gracilis (2n = 4), a study has been made of the growth of its stem tissue in culture. Although growth may occur on a basal medium supplemented in various ways, it was stimulated for present purposes by the use of a basal medium containing casein hydrolysate, coconut milk (2-10% by volume) and naphthaleneacetic acid (0.5 p.p.m.). A definite synergism between coconut milk and naphthaleneacetic acid was demonstrated. The general form of the colonies so obtained responds to the composition of the medium, and certain effects of pigmentation indicate that the biochemistry of the cultured tissue is also a function of the conditions. The Haplopappus cultures were maintained in liquid culture either in the form of free cells or of the small cell clusters to which they readily gave rise. The form of typical cells and cell clusters is described, and stress is laid upon the range of growth forms that are encountered. Variations in suspensions of cells, or small cell clusters, may be investigated by the application of simple microbiological techniques. Haplopappus gracilis is thus a useful material for the further study of growth and morphogenesis by tissue culture techniques. MODERN developments in the technique of culturing explanted tissues, as well as subcultures and freely suspended cells derived from these, have greatly increased the range of the plant tissueculture technique. Improved methods have increased the prospects for the use of cell and tissue cultures and have also increased the range of their responses which are now open to investigation. In particular, the prospects for the use of cell and tissue cultures in the investigation of morphogenesis have been enhanced. Among the developments which have proved to be significant in this laboratory are the following: (1) the recognition that coconut milk, and other similar fluids, may reactivate quiescent cells and cause their renewed growth by rapid cell division at previously unprecedented rates; (2) the recognition that many tissues, that are otherwise recalcitrant, respond to synergistic combinations of coconut milk with a variety of synthetic growth regulators of the auxin type, and to media that are supplemented with casein hydrolysate as a source of reduced nitrogenous compounds; (3) the development of procedures for the controlled and aseptic growth of tissue cultures in liquid media; and (4) the recognition that freely suspended cells, disassociated from tissue cultured in liquid, are also cap1 Received for publication November 28, 1960. This work forms part of a continuing program of research which is being supported by grants to one of us (F.C.S.) from the National Cancer Institute, National Institutes of Health, Department of Health, Education, and Welfare. The grant C-1359 (C9) made possible the assistance of Mrs. Marion 0. Mapes, for whose help with the photography the authors are especially grateful. During the course of the work, one of .us (L.M.B.) held a predoctoral fellowship under the cooperative arrangements between Cornell University and the National Science Foundation. able of growth and may then reveal other hitherto undisclosed capacities for organization and develop-