SummaryWe have developed a strategy for the optimization of plant cell suspension culture media using a combination of fractional factorial designs (FFDs) and response surface methodology (RSM). This sequential approach was applied to transformed tobacco BY-2 cells secreting a human antibody (M12) into the culture medium, in an effort to maximize yields. We found that the nutrients KNO 3 , NH 4 NO 3 and CaCl 2 and the hormones 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (BAP) had the most significant impact on antibody accumulation. The factorial screening revealed strong interactions within the nutrients group (KNO 3 , NH 4 NO 3 and CaCl 2 ) and also individually between 2,4-D and three other components (KNO 3 , NH 4 NO 3 and BAP). The RSM design resulted in a fivefold increase in the antibody concentration after 5 days and a twofold reduction in the packed cell volume (PCV). Longer cultivation in the optimized medium led to the further accumulation of antibody M12 in the culture medium (up to 107 lg/mL, day 10). Because the packed cell volume was reduced in the optimized medium, this enhanced the overall yield by 20-fold (day 7) and 31-fold (day 10) compared to the conventional MS medium.

Callus and hairy root cultures of Linum leonii were established. The genetic transformation in hairy roots was proven by PCR analysis, which showed integration of rol A and rol C genes into the plant genome. Calli and hairy roots accumulate the arylnaphthalene lignan justicidin B as a major constituent. Hairy roots produce 5-fold higher yields of justicidin B (10.8 mg g(-1) DW) compared to calli. Justicidin B shows strong cytotoxicity on the chronic myeloid leukemia LAMA-8 and K-562 cell lines and on the chronic lymphoid leukemia SKW-3 cell line with IC(50) values of 1.11, 6.08, and 1.62 microM, respectively. Apoptotic properties of justicidin B are reported for the first time.

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