PHOTOAUTOTROPHIC GROWTH OF SOYBEAN CELLS IN SUSPENSION-CULTURE .2. OPTIMIZATION OF CULTURE-MEDIUM AND CONDITIONS

We have attempted to optimize the conditions under which a photoautotrophic soybean suspension culture line (SB-P; Horn et al. 1983) is grown. Magnesium, phosphate, and calcium concentrations were varied individually from one-tenth to five times the normal level found in the Murashige and Skoog (1962) recipe. After two subcultures, only phosphate at one-tenth the normal level caused the cells to show a substantial reduction in fresh and dry weight increase and chlorophyll level. Nitrate and ammonium levels were inversely varied in 20 millimolar increments of potassium nitrate and ammonium chloride. Neither N-source alone could support growth through two subcultures. A ratio of 40 millimolar potassium nitrate to 20 millimolar ammonium gave significantly better fresh and dry weight increases than did a ratio of 20:40 or 30:30 but the chlorophyll level was unchanged. The minor salts as a group resulted in a small improvement in growth when provided at twice the normal level. Indole-3-acetic acid at five milligrams per liter resulted in significantly better fresh and dry weight increases than did alpha-naphthaleneacetic acid at any level but the final chlorophyll level was not changed. There was no correlation between growth and kinetin level and this resulted in the discovery that SB-P cells are cytokinin-autotrophic, as are heterotrophic SB cells, with regard to both growth and greening ability. Growing SB-P cells under a 14 h: 10 h day: night photoperiod resulted in a slow but inevitable death. Increasing the carbon dioxide level to 10% for four weeks gave no increase in SB-P cell growth or chlorophyll level, but SB-P cells would not grow with carbon dioxide levels below 0.4%. The results clearly show that SB-P cells, despite their tenuous existence, are capable of adapting to a wide range of culture conditions. A simplified and improved culture medium for photoautotrophic cultures is given.