[Objective] This study aimed to explore the protoplast preparation and re- generation conditions of Tremella aurantialba. [Method] Optimal combination of five factors affecting the protoplast preparation and regenerat...[Objective] This study aimed to explore the protoplast preparation and re- generation conditions of Tremella aurantialba. [Method] Optimal combination of five factors affecting the protoplast preparation and regeneration of Tremella aurantialba was selected by using orthogonal experiment, including enzyme system, culture age, enzymolysis temperature, enzymolysis duration and osmotic stabilizer. [Result] The results showed that there were three release modes of Tremella aurantialba proto- plasts： release from top in the early period of enzymolysis, release from the side and release in situ. The optimal protoplast preparation condition was selecting mycelium cultured in liquid medium for 3 d, for enzymolysis in mixed enzyme solu- tion containing 1% of cellulase ＋1% of snailase ＋1% of lywallzyme at 36 ℃; for 4 h, with 0.6 mol/L sucrose as osmotic stabilizer, and the obtained preparation rate had reached 1.76 ×10^7 protoplasts/ml. The optimal regeneration condition was using mycelium cultured in liquid medium for 5 d, for enzymolysis in enzyme solution con- taining 1.5% of lywallzyme at 33 ℃ for 3 h, with 0.6 mol/L sucrose as osmotic sta- bilizer, and the regeneration rate had reached 0.22%. [Conclusion] This study provid- ed valuable support for protoplast fusion, ultraviolet mutation breeding and genetic engineering research.展开更多
文摘[Objective] This study aimed to explore the protoplast preparation and re- generation conditions of Tremella aurantialba. [Method] Optimal combination of five factors affecting the protoplast preparation and regeneration of Tremella aurantialba was selected by using orthogonal experiment, including enzyme system, culture age, enzymolysis temperature, enzymolysis duration and osmotic stabilizer. [Result] The results showed that there were three release modes of Tremella aurantialba proto- plasts： release from top in the early period of enzymolysis, release from the side and release in situ. The optimal protoplast preparation condition was selecting mycelium cultured in liquid medium for 3 d, for enzymolysis in mixed enzyme solu- tion containing 1% of cellulase ＋1% of snailase ＋1% of lywallzyme at 36 ℃; for 4 h, with 0.6 mol/L sucrose as osmotic stabilizer, and the obtained preparation rate had reached 1.76 ×10^7 protoplasts/ml. The optimal regeneration condition was using mycelium cultured in liquid medium for 5 d, for enzymolysis in enzyme solution con- taining 1.5% of lywallzyme at 33 ℃ for 3 h, with 0.6 mol/L sucrose as osmotic sta- bilizer, and the regeneration rate had reached 0.22%. [Conclusion] This study provid- ed valuable support for protoplast fusion, ultraviolet mutation breeding and genetic engineering research.
