Decrease in the regeneration potential of long-term cell suspension cultures of Lilium formosanum Wallace and its...

Cell suspension cultures of Lilium formosanum Wallace were initiated from bulb scale-derived calli and subcultured every 2 weeks using a medium containing 5 µM 4-amino-3,5,6-trichloropicolinic acid (picloram). Almost all cell clumps from the suspension cultures developed numerous somatic embryos following their transfer onto a plant growth regulator-free medium, while they vigorously produced shoot buds on media containing 0.5 or 5 µM 6-benzyladenine (BA). The high regeneration potential on a plant growth regulator-free medium was maintained for up to 54 months, but it gradually decreased thereafter, and only a few adventitious shoots and embryos were obtained from 75-month-old cultures. For restoring the regeneration potential of these cultures, various treatments with plant growth regulators were applied, among which about 10-fold increases in the number of regenerated shoot buds were obtained with 0.5 or 5 µM 2,3,5-triiodobenzoic acid (TIBA) in combination with 0.5 or 5 µM BA or N-(1,2,3-thiadiazol-5-yl)-N'-phenylurea (thidiazuron). Only shoot buds were produced from the cell clumps cultured on TIBA-containing media, and these shoot buds developed into complete plantlets after they were excised from the calli and transferred to a plant growth regulator-free medium.