POLYAMINE METABOLISM ASSOCIATED WITH GROWTH AND EMBRYOGENIC POTENTIAL OF RICE

To determine whether polyamines are involved in somatic embryogenesis in a monocot, levels of polyamines were determined using HPLC during embryogenic callus induction and during cell-culture growth of rice (Oryza sativa L.). Free putrescine accumulated ten-fold in cells during callus induction and growth using conditions optimized for subsequent plant regeneration. Free and conjugated pools of putrescine, spermidine, and spermine in cell suspension cultures exhibited a regular, proportional pattern of increase and decline during each subculture cycle. Polyamine biosynthesis inhibitors, difluoromethylarginine (DFMA) and difluoromethylornithine (DFMO), were used to determine the importance of de novo putrescine biosynthesis for explant dedifferentiation and growth of embryogenic rice callus. Callus was induced in the presence of 2,4-D and either of these inhibitors. However, DFMA (10 mM) suppressed cell suspension culture growth and plant regeneration from callus. These effects were reversible with exogenous agmatine. DFMO inhibited cell suspension culture growth only in the absence of exogenous 2,4-D and had no effect on plant regeneration from callus. A comparison of the morphogenesis of cell suspensions cultured in the presence of DFMA or DFMO revealed that these inhibitors differ in their impact on cell differentiation, proliferation of compact meristematic cell clusters, starch metabolism, and cell cluster friability. We conclude that polyamine metabolism via arginine decarboxylase influences the embryogenic potential of in vitro rice cultures.