含抗菌肽CC31毕赤酵母工程菌培养基及其培养条件的优化

目的筛选毕赤酵母工程菌培养基的主要组分,优化工程菌培养条件。方法采用毕赤酵母工程菌CC31发酵培养抗菌肽CC31,选取基础盐培养基(basic salt medium,BSM)为初始培养基,利用单因素试验、正交试验设计对培养基的碳源、氮源、酵母营养物进行筛选;选取最适培养基对菌种接种量、培养基初始pH值、培养温度进行培养条件优化。结果经鉴定,抗菌肽CC31相对分子质量为13 620;初始BSM培养基添加组分中碳源为3%甘油和3%葡萄糖,酵母营养物为1%酵母浸出物,氮源为1. 5%硫酸铵;在10%接种量、培养基初始pH值为6,培养温度为30℃的培养条件下,获得的抗菌肽CC31蛋白浓度可达82 mg/L。结论优化了毕赤酵母工程菌培养基组分及培养条件,获得高表达量的重组蛋白,为工业化高密度发酵提供了实验依据。

Optimization of culture medium and culture condition for recombinant Pichia pastoris containing CC31

Objective To screen the main components of medium and optimize the condition for culture of recombinant Pichia pastoris. Methods Antibacterial peptide CC31 was subjected to fermentation culture with recombinant P. pastoris CC31 in basic salt medium(BSM). The carbon source,nitrogen source and yeast nutrient in BSM were screened by single factor experiment and orthogonal experiment. The culture conditions in the optimal medium,including the inoculation quantity,initial pH value and temperature,were optimized. Results The relative molecular mass of antibacterial peptide CC31 was 13 620. The optimal carbon source added into BSM was 3% glycerin and 3% glucose,while the optimal nitrogen was 1. 5% ammonium sulphate,and the yeast nutrient was 1% yeast extract. The optimal inoculation quantity,initial pH value and temperature were 10%,6 and 30 ℃ respectively,under which the cultured CC31 reached 82 mg/L.Conclusion The medium component and culture condition for recombinant P. pastoris were optimized,and recombinant CC31 was highly expressed,which provided an experimental basis for industrial high density fermentation of recombinant P. pastoris.