Establishment and characterization of photoautotrophic protoplast-derived cultures ofNicotiana plumbaginifolia.

A procedure is described for the rapid establishment of photoautotrophic protoplast-derived cultures ofNicotiana plumbaginifolia. Photoautotrophic growth was induced by lowering the glucose concentration to 2.5 g.l(-1) in the protoplast culture medium and by omitting glucose from the subsequent dilution medium. Four week-old highly viable suspensions were plated on an agar-medium without glucose in unsealed Petri dishes and kept in illuminated chambers flushed with 0.05 % or 2 % CO2. Air-grown calli had net photosynthesis rates of 1.8 and 17 μmoles CO2.g(-1) fresh wt.h(-1) in air at 0.034 % CO2 and in air enriched with 1 % CO2, respectively. Calli grown in 2 % CO2 exhibited lower rates of net photosynthesis at the two CO2 concentrations tested (0 and 7.5 μmoles CO2.g(-1) fresh wt.h(-1), respectively). The contribution of photosynthesis to growth was estimated to be 80 % in air-grown calli and more than 90 % in calli grown in 2 % CO2. The suitability of this photoautotrophic culture procedure is discussed with regard to the screening of photosynthetic mutants or transformants from protoplasts.