摘要
[目的]研究来源于苏云金芽孢杆菌的几丁质酶基因chiA在枯草芽孢杆菌中的表达情况。[方法]以苏云金芽孢杆菌染色体DNA为模板,PCR扩增获取几丁质酶基因chiA,将其与枯草芽孢杆菌表达载体pHSG连接,构建重组菌,经IPTG诱导后,检测培养液中的几丁质酶活性。[结果]扩增得到几丁质酶基因chiA大小为2.5 kb。构建的重组菌对底物[4-MU-(GlcNAc)3]显示出一定的水解活性,培养液酶活约为2.8 U/ml,而pHSG空质粒转化子在同样条件下其培养液没有明显酶活。该重组酶的最适pH值为6.5,最适反应温度为50℃,与苏云金芽孢杆菌自身产生的几丁质酶性质一致。[结论]几丁质酶基因chiA能在枯草芽孢杆菌中成功表达,表达产物可成功分泌到细胞外。
[ Objective ] The aim was to study the expression of the chitinase gene chiA from Bacillus thuringiensis in Bacillus subtilis. [ Method] The chitinase gene chiA was amplified from the chromosome DNA of B. thuringiensis by PCR, then connected with pHSG expressive vector, and then constructed recombinant strain. The chitinase activity of culture medium was detected after IPTG induction. [ Result] The chitinase gene chiA of 2.5 kb was obtained by amplification. The constructed recombinant strain exhibited certain hydrolytic activity on the substrate [4-MU-（ GIcNAc）3 ] and the enzyme activity of culture medium was about 2.8 U/ml, while the transformant of pHSG empty plasmid showed no obvious enzyme activity in its culture medium under the same condition. The optimum pH value of the recombinase was 6.5, its optimum reaction temperature was 50 ℃ and its characteristic of chitinase was consistent with that of chitinase produced by the B. thuringiensis itself. [ Conclusion] The chitinase gene chiA could express in B. subtilis successfully and the expressive production could excrete to extracellular successfully.
出处
《安徽农业科学》
CAS
北大核心
2008年第26期11242-11244,11247,共4页
Journal of Anhui Agricultural Sciences
基金
国家高技术研究发展计划(863计划
2006AA10Z307)资助项目
