摘要
目的:建立一种使基因工程中重组蛋白质高效表达并提高可溶性比率的新方法。方法:2002-09/2005-03在新乡医学院分子生物研究室用不含葡萄糖的M9ZB培养基分别培养转化过的含Tac启动子或T7启动子的工程菌,采用终浓度为0.02mmol/L的IPTG及5mmol/L的乳糖进行诱导。采用SDS-PAGE及岛津薄层扫描分析仪进行表达效率检测、蛋白可溶性的扫描分析。结果:用常规方法(1mmol/LIPTG,LB培养基)诱导,重组基因的表达效率不高,表达量占菌体总蛋白25%,且可溶性产物比例较低,占表达蛋白的20%,采用0.02mmol/L的IPTG及5mmol/L乳糖诱导后重组蛋白表达效率占菌体总蛋白的65%,可溶性产物比例亦大幅升高占表达蛋白90%～93%,此方法没有选择性,对T7启动子表达系统和Tac启动子表达系统均有效。结论:采用0.02mmol/L的IPTG及5mmol/L乳糖诱导后,不仅使含Tac启动子或T7启动子的表达系统中重组蛋白表达增加而且产物可溶性升高。
AIM： To establish a method which enhances protein expression and solubility of recombinant protein in gone engineering.
METHODS： This experiment was conducted at the Molecular Biology Laboratory,, Xinxiang Medical College from September 2002 to March 2005. M9ZB culture medium without glucose was used to culture transformed engineering bacteria containing Tac and T7 promotore. Isopropylthiogalactoside （IPTG） with final concentration of 0.02 mmol/L and lactose with final concentration of 5 mmol/L were used. SDS-PAGE and scanning monitor were used to detect expression efficiency, and analyze the solubility of protein.
RESULTS： Routine method （1 mmol/L IPTG,LB culture medium） was used to culture, and the expression efficiency of recombinant gane was not high. Expression was 25% of the thallus total protein, and the proportion of soluble products was very low, 20% of expression protein. After induction of 0.02 mmol/L IPTG and 5 mmol/L lactose, expression efficiency of recombinant protein was 65% of the thallus total protein, the proportion of soluble products was increased greatly, being 90%-93% of expression protein. This method was useful not only to the Tac promoter but also to the I＂7 promoter.
CONCLUSION： Induction of 0.02 mmol/L IPTG and 5 mmol/L lactose not only increases the recombinant protein expression of expression system containing Tac or T7 promotor but also enhances the solubility of products.
出处
《中国临床康复》
CSCD
北大核心
2006年第29期92-93,共2页
Chinese Journal of Clinical Rehabilitation
基金
河南省教委资助项目(20011800002)~~
