ABSTRACIActive polyamine biosynthesis occurs in the embryonic axis, but not in the cotyledons, during germination of Glyciac max (L.) further elucidated. To understand the potential roles of polyamines in cellular metabolism, it is necesary to clarify whether changes in the content of polyamines in general or a specific polyamine is causally linked to, or merely a consequence of, plant growth and development. In this study, polyamine levels and complexities were determined in the cotyledon, hypocotyl hook, hypocotyl, and radicle of soybean (Glycine mnax L. Merr.) cv Williams during and after seed germination. Experiments were also carried out to characterize Cad biosynthesis in the embryonic axis during seed germination and growth ofthe young seedling.
MATERIAIS AND METHODS Plant Material. Seeds of the soybean (Glycine max L. Menf.) cv Williams harvested in 1982 from Spindletop ExperimentalFarm of the University of Kentucky were used in this study. Unless otherwise noted, seeds were allowed to germinate at 23°C in the dark on water-moistened Anchor seed germination paper (Anchor Paper Co., St. Paul, MN). Samples were taken at intervals throughout germination and the cotyledons and germinated embryonic axes were manually separated. Unless otherwise stated, each of the embryonic axes was divided into three parts (radicle, hypocotyl, and hypocotyl hook) for determination of fresh weight, dry weight, and polyamine content. The axes could be clearly divided into the above three distinguishable parts by 2 d after the seeds were placed on the water-moistened germination paper. The axes from dry seed or seed that had been on the germination paper for up to 1.5 d were divided by length with one-quarter the length from the apex desgnated as the radicle, the following one-quarter length as the hypocotyl, and the remaining part as the hypocotyl hook. The separation of the hypocotyl hook from the hypocotyl is based on the observation that the hypocotyl hook portion turns greenish in color within hours after exposure to light ofdetached embryonic axes cultured in a nutrient agar medium (20). The hook portion in darkgerminated seedlings is yellowish in color.Analysis of Polyamine Content. Routinely, the fresh tissue derived from 30 embryonic axes (300-800 mg fresh weight) per assay was homogenized in 5 ml ofcold 5% HC104 with a Polytron tissue homogenizer. Polyamines in the acid-soluble and -insoluble fractions were qualitatively and quantitatively determined by a HPLC system as described previously (10). In some experiments, 5 to 15 seedlings (after 3 d of imbibition) were used per assay and the seedlings were homogenized as described. Authentic polyamines were used for calibration of polyamine content in the extracts.