新生隐球菌原生质体形成条件的研究

目的：研究新生隐球原生质体形成的条件。方法：应用不同的酶（溶壁酶和NovoZyme234）、在不同的酶浓度（1、3、5mg/ml）、pH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0.8mol/L的山梨醇为渗透压稳定剂，pH6.0，加入5mmol/L DTT和3mg/ml NovoZyme234，孵育75min，在此条件下，原生质体形成率是98.1%。溶壁酶与NovoZyme234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、pH值和渗透压稳定剂。

Study on experimental conditions for production of Cryptococcus neoformans protoplasts

Objective: To establish conditions for the protoplasts formation of C.neoformans . Methods: Protoplasts from C.neoformans were prepared by various enzymes (lywallzyme and NovoZyme234), concentration of enzyme (1,3,5 mg/ml), pH values (pH 4,5,6,7,8) and osmotic stabilizers (sucrose,mannitol and sorbitol). The rate of protoplasts formation were counted.Results: The early exponential phase cells were easy to form protoplasts, under the action of pH 6.0 and 5 mmol/L DTT, 0.8 mol/L sorbitol used as osmotic stabilizer, NovoZyme234 at the concentration of 3 mg/ml was most effective for the protoplasts yield. The ratio of protoplasts formation was 98.1% in 75 min. The results of lywallzyme were similar to NovoZyme234. Conclusion:Key factors for success in isolation of protoplasts are the availability of suitable lytic enzymes,pH value and osmotic stabilizer. [