铜绿假单胞菌脂肪酶Lipase基因的原核表达  

Prokaryotic Expression of Pseudomonas aeruginosa Lipase Gene

作　　者：张煜星[1,2,3] 武寒雪[3] 祝建波[3] 刘焕[4] 周鹏[1,2] 

机构地区：[1]海南大学农学院,海南儋州571737 [2]中国热带农业科学院热带生物技术研究所国家重点实验室,海南海口571101 [3]石河子大学生命科学学院农业生物技术重点实验室,新疆石河子832003 [4]新疆农业科学院微生物应用研究所,新疆乌鲁木齐830091

出　　处：《安徽农业科学》2008年第35期15384-15385,15388,共3页Journal of Anhui Agricultural Sciences

基　　金：国家"863"子项目"特殊生境植物资源的开发利用技术"(2007AA021401);兵团博士基金项目"转基因育种技术研究"(2006JC07)

摘　　要：[目的]对铜绿假单胞菌脂肪酶Lipase基因进行原核表达。[方法]利用PCR方法从铜绿假单胞菌基因组DNA中扩增得到脂肪酶基因,测定其核苷酸序列,利用基因重组技术构建脂肪酶基因的原核表达载体,加IPTG至终浓度为1.0 mmol/L诱导蛋白表达4 h,并进行SDS-PAGE电泳。[结果]从铜绿假单胞菌中克隆的脂肪酶基因成熟肽的序列,与NCBI上所递交的铜绿假单胞菌脂肪酶序列同源性很高,达99.36%。成功构建了脂肪酶基因的原核表达载体pET32a-Lip,进一步SDS-PAGE电泳结果显示目的基因得到高效表达。[结论]克隆的铜绿假单胞菌脂肪酶带有自身的信号肽,也可以在大肠杆菌中正常表达,可以用于进一步的研究。[ Objective] The aim of this study was to investigate the prokaryotie expression of pseudomonas aeruginosa lipase gene. [ Method] Lipase gene was amplified by PCR from the genome DNA of pseudomonas aeruginosa, and its nueleotide sequence was determined. The prokaryotic expression vector of Lipase gene was constructed by the gene recombination technique. The protein expression was induced for 4 hours by IPTG with the final concentration of 1.0 mmol/L, and then SDS-PAGE electrophoresis was analyzed. [ Result] The sequence of mature peptides in lipase gene cloned from pseudomonas aeruginosa had a 99.36% homology with that of pseudomonas aeruginosa lipase submitted in NCBI, so the prokaryotic expression vector of lipase gene pET32a-Lip was successfully constructed. Furthermore, the results of SDS-PAGE electrophoresis showed that the target gene was expressed highly and effectively. [ Conclusion] The cloned pseudomonas aeruginosa lipase with its signal peptide can be normally expressed in Escherichia coli and also used for further study.

关 键 词：铜绿假单胞菌 脂肪酶 原核表达 

分 类 号：S188[农业科学—农业基础科学]