title: Plant regeneration from protoplasts isolated from callus ofGentiana crassicaulis.
abstract: Fast growing calli induced from hypocotyl segments ofGentiana crassicaulis were used for preparation of protoplasts. High yields of viable protoplasts were produced in an enzyme solution containing 1-2% cellulase, I% pecfinase, and 0.5% Hemicellulase. Protoplasts were cultured in KM8P medium containing 1 mg/l 2,4-D, 0.5 mg/l 6BA, 500 mg/l LH, 0.5 M glucose and 0.1 M mannitol by the solid-liquid dual layer culture method. First division occurred within 4-5 days of culture at a frequency of 17.8%. Sustained divisions led to callus formation. Periodically diluting the cultures with freshly prepared liquid medium containing 1% glucose was critical for colony formation. Protocolonies about 2 mm in size were transferred onto MS medium supplemented with 3 mg/l ZT, 2 mg/l 6BA, 1 mg/l GA3, 1 mg/l NAA and 6% sucrose to obtain embryogenic calli. Plantlets were regenerated via somatic embryogenesis at high frequency on hormone-free MS Medium.
authors.affiliation: Biology Department, Lanzhou University, 730000, Lanzhou, Gansu, People's Republic of China.
authors.forename: Y L
authors.initials: YL
authors.lastname: Meng
authors.forename: Y P
authors.initials: YP
authors.lastname: Gao
authors.forename: J F
authors.initials: JF
authors.lastname: Jia