Preparation of protoplasts from Chlorella protothecoides |
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Authors: | Yongzhong Lu Renqiu Kong Liwei Hu |
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Affiliation: | (1) Biology Department, Qingdao University of Science and Technology, Qingdao, 266042, People’s Republic of China;(2) State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, 430071, People’s Republic of China |
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Abstract: | This paper describes an enzymatic method for yielding protoplasts from the microalga Chlorella protothecoides. Four kinds of commercially available enzymes were tested. The enzymatic digestion was optimal with 2% cellulase R-10 and 1% snailase prepared in 25 mM Tris buffer (pH 6.0) containing 0.6 M D-mannitol, and the protoplast density could reach the peak after treatment at 30°C for 16 h. Nearly all liberated protoplasts were green in the presence of 0.01% phenosafranin, indicating their high viability. The regeneration rate was about 70% when 0.6 M D-mannitol was used as an osmotic stabilizer in the regeneration medium. This protocol will find useful applications in genetic studies of this algal species. |
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