The role of N metabolism in organogenesis and growth was studied using tobacco pith callus. Callus was cultured on a solid medium containing 10 μM (1.75 mg/l) IAA and 2 μM (0.43 mg/l) kinetin for 56 days. In the growth experiment, ratios of NH4 +-N to NO3 --N (0:60, 20:40, 30:30, 40:20 and 60:0 mM) were supplied by (NH4)2 SO4 and KNO3. Callus and media were analyzed for inorganic N. Callus supported by 30:30 and 40:20 media removed the highest amounts of NH4 +-N and NO3 --N from the media and exhibited organogenesis. Final dry weight was greatest in callus supported by the 30:30 medium. In the organogenesis experiment, the transfer history of the inoculum source affected N uptake, organogenesis and growth. Inorganic N was supplied by NH4NO3 and KNO3 -. The net uptake of NH4 +-N and NO3 --N was lower in shoot-forming than in root-forming and non-organogenic callus subculture from 7-day-old stock cultures. The final pH of the medium supporting shoot-forming callus was lowest. Growth, on a dry weight basis, was lowest in shoot-forming callus. Callus subculture from 60-day-old stock cultures formed no shoots.
Victoria E. Rudolph and David W. Burger
Suzanne M.D. Rogers, Victoria E. Rudolph and Kalyani Dias
A suspension culture of Eucalyptus tereticornis was initiated from callus and grown for 7 months under indirect light in a Murashige and Skoog (1962) basal medium containing 3% glucose and 1 mg/l 2,4-D. Glucose was used, instead of sucrose, as it reduced production of phenolic-like compounds. The inoculum size for maximum cell yield was determined. Cells (0.1, 0.2, 0.5, 1.0, 2.5 and 5.0 g fresh weight) were cultured in basal medium for 14 days. Maximum fresh weight (mean 11.8 g) was attained from samples inoculated with at least 1.0 g of cells. Largest dry weight (mean 608 mg) occurred following, inoculation with at least 0.5 g fresh weight of cells. Inoculation with 0.5 g of cells resulted in the most rapid fresh weight doubling time (3.4 days).
After 17 months of culture, cells were grown in basal medium or m basal medium supplemented with 1 mg/l kinetin, under continuous, direct light. Growth, based on fresh and dry weight increases, was measured over the 2-week subculture period. Growth of cells was similar in both media. The cells' chlorophyll content remained low. Fresh weight doubling time averaged 3.8 days.