食品科学 ›› 2013, Vol. 34 ›› Issue (9): 140-142.doi: 10.7506/spkx1002-6630-201309029

• 生物工程 • 上一篇    下一篇

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   

  1. 贵州大学 生命科学学院
  • 收稿日期:2012-09-20 修回日期:2013-03-14 出版日期:2013-05-15 发布日期:2013-05-07
  • 通讯作者: 吴拥军 E-mail:wyjbio@163.com
  • 基金资助:

    国家自然科学基金地区基金项目(31260394);贵阳市科技计划项目(筑科工合同字[2010]第1-68号)

Cloning and Expression of Glutaminase Gene glsA2 in Prokaryotic System

LU Biao,WU Yong-jun*,WU Yu-jun,LUO Xi,LIU Yan-min   

  • Received:2012-09-20 Revised:2013-03-14 Online:2013-05-15 Published:2013-05-07
  • Contact: WU Yong-jun E-mail:wyjbio@163.com

摘要:

为验证谷氨酰胺酶基因glsA2的酶活力,以pET-32a为表达载体构建原核表达重组质粒pET32a-glsA2,转化表达菌株E.coli BL21(DE3)。从异丙基-β-D-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glsA2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。

关键词: 谷氨酰胺酶, glsA, 原核表达, IPTG诱导

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: glutaminase;glsA;prokaryotic expression;isopropyl &beta, -D-thiogalactopyranoside (IPTG) induction

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