谷氨酰胺酶基因原核表达载体的构建与表达 - 食品科学.pdf

140 2013, Vol.34, No.09 食品科学 ※生物工程 谷氨酰胺酶基因原核表达载体的构建与表达 * 卢 彪，吴拥军 ，吴玉俊，罗 熹，刘艳敏 (贵州大学生命科学学院，贵州 贵阳 550025) 摘  要： glsA 的酶活力，以pET-32a为表达载体构建原核表达重组质粒pET32a-glsA ，转化表 为验证谷氨酰胺酶基因 2 2 E.coli β D glsA 达菌株 BL21(DE3) 。从异丙基- - -硫代吡喃半乳糖苷(IPTG) 的诱导浓度及诱导表达时间两方面对重组菌株 2 基因的表达系统进行优化。结果显示，0.1mmol/L IPTG诱导6h ，谷氨酰胺酶活力达到608.2U/µg，约为对照的15倍。 glsA 关键词：谷氨酰胺酶； ；原核表达；IPTG诱导 Cloning and Expression of Glutaminase Gene glsA2 in Prokaryotic System * LU Biao，WU Yong-jun ，WU Yu-jun ，LUO Xi ，LIU Yan-min (College of Life Sciences, Guizhou University, Guiyang 550025, China) Abstract ：To test the activity of glutaminase from Bacillus subtilis BJ3-2, the gene glsA2 was cloned into prokaryotic expression vector pET-32. The recombinant plasmid pET32a-glsA2 was then transformed into E.coli BL21 competent cells. Optimum IPTG concentration and induction time for gene glsA2 expression were optimized. The result showed that the expression was optimized with proper inducing conditions of 0.1 mmol/L IPTG for 6 h. Under these conditions, glutaminase activity in supernatant of cell lysate reached 608.2 U/μg protein, which was 15 times higher than that of control. Key words glsA β-D ：glutaminase ； ；prokaryotic expression ；isopropyl -thiogalactopyranoside (IPTG) induction 中图分类号：Q939.97 文献标志码：A