关键词: 磷脂酰肌醇特异性磷脂酶C, 克隆表达, 乳酸乳球菌, GPI锚定蛋白

Abstract: Based on the codon bias of Lactococcus lactis， phosphatidylinositolspecific phospholipase C gene from Bacillus cereus was optimized and then synthesized for protein expression. The cloned gene was inserted into Escherichia coliLactococcus lactis shuttle vector pAMJ399， and then transformed to L. lactis cells by electroporation to induce expression. The result of SDSPAGE showed that the recombinant protein was secreted extracellular in the form of soluble proteins， and its molecular weight was about 35 ku， which was consistent with the expected protein size. Meanwhile， the recombinant protein showed significant enzyme activity on PIListeria chromogenic plate. The results indicated that phosphatidylinositolspecific phospholipase C （PIPLC） was successfully expressed in L. lactis. The growth condition was optimized as follows: 2% inoculation amount; GM17 medium with 1% erythromycin; 32 ℃. After 24 h static culture under above condition， it could produce 1.092 μg·mL-1 PIPLC in the supernatant of culture medium.

Key words: phosphatidylinositolspecific phospholipase C, cloning and expression, Lactococcus lactis, GPIanchored proteins