蜗牛酶高效制备白色念珠菌原生质体的研究

Preparing high quality protoplast of Candida albicans by snailase

为了寻求以蜗牛酶经济高效的白色念珠菌原生质体的制备方法,以白色念珠菌为材料,研究了菌龄、预处理剂、酶浓度、作用时间、高渗稳定剂等因素对原生质体制备率和再生率的影响。结果表明,以蜗牛酶制备白色念珠菌原生质体的最佳菌龄是培养10～14 h的菌体,最佳预处理剂配方为50 mmol/L Tris(pH8.0)、50 mmol/L EDTA和5%巯基乙醇,1%蜗牛酶在37℃酶解1 h原生质体制备率可达90%以上,此原生质体采用夹层平板培养法在0.7 mol/L NaCl高渗培养基中培养,再生率最高可达95%。本研究表明单独用蜗牛酶高效制备高质量的白色念珠菌原生质是可行的。

To look for methods of preparing high quality protoplast of Candida albican at high rates, pretreating agent, cell age, osmotic stabilizing agent, incubation temperature and time, as well as enzyme concentration were taken into consideration in the cause of preparing and regenerating protoplasts from Candida albicans. The results indicated that the optimum condition for protoplast preparation and regerneration were 1% snailase at 37℃, and the yeast inoculated for 10-14 h and pretreaed with 5% of mercaptoethanol in the presence of 50 mmol/L EDTA and 50 mmol/L Tris base （pH8.0）. At the above condition, the protoplast production could surpass 90% within 1 h. The regeneration rate of the obtained protoplasts could reach 95% after bilayer-cultivated in Sabouraud＇s medium containing 0.7 mol/L NaCl. It is feasible to prepare high quality C. albicans protoplast by snailase only.