Plant regeneration from protoplasts isolated from callus ofGentiana crassicaulis |
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Authors: | Yu-ling Meng Yun-peng Gao Jing-fen Jia |
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Institution: | (1) Biology Department, Lanzhou University, 730000 Lanzhou, Gansu, People's Republic of China |
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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.Abbreviations 6BA
6-benzylaminopurine
- NAA
naphthaleneacetic acid
- 2,4-D
2,4 - dichlorophenoxyacetic acid
- ZT
zeatin
- GA3
gibberellic acid
- LH
lactalbumin hydrolysate
- MES
2-(N-morpholino)-ethane sulfonic acid
- MS
Murashige & Skoog's medium(1962) |
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Keywords: | Gentiana crassicaulis Duthie ex Burk protoplasts callus plant regeneration |
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