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目的 :研究新生隐球菌原生质体形成的条件。 方法 :应用不同的酶 (溶壁酶和 Novo Zym e2 34 )、在不同的酶浓度 (1、3、5 m g/ml)、p H值 (4、5、6、7、8)以及加入不同的渗透压稳定剂 (蔗糖、甘露醇和山梨醇 )的条件下 ,分别制备原生质体并计算原生质体形成率。 结果 :对数生长早期的细胞较易形成原生质体 ,原生质体形成的最佳条件是以 0 .8m ol/L的山梨醇为渗透压稳定剂 ,p H 6 .0 ,加入 5 m mol/L DTT和 3mg/ml Novo Zym e2 34 ,孵育 75 m in,在此条件下 ,原生质体形成率是 98.1%。溶壁酶与 Novo Zym e2 34的结果相似。 结论 :原生质体分离的关键因素是合适的溶菌酶、p H值和渗透压稳定剂
Objective: To study the conditions of Cryptococcus neoformans protoplast formation. METHODS: Different enzymes (Lysozyme and Novo Zym e2 34) were used at different enzyme concentrations (1, 3, 5 mg / ml), p H values (4, 5, 6, 7, 8) Protoplasts were prepared separately and the protoplast formation rate was calculated under the conditions of osmotic pressure stabilizers (sucrose, mannitol and sorbitol). RESULTS: Protoplasts were easier to form protoplasts in early logarithmic growth phase. The optimum conditions for protoplast formation were as follows: 0 .8 mol / L sorbitol as osmotic pressure stabilizer, p H 6 .0, 5 mol / L DTT and 3 mg / ml Novo Zym e234 for 75 mins. Under these conditions, the protoplast formation rate was 98.1%. Lysozyme was similar to Novo Zym e234. Conclusions: The key factor in protoplast isolation is the appropriate lysozyme, p H value and osmolarity stabilizer