氮离子诱变筛选内切葡聚糖酶高产菌株及其基因突变研究  

Studies on Endoglucanase Activity and Gene Mutation of Bacillus subtilis Induced by Nitrogen Ion Irradiation

作　　者：吕杰[1,2] 金湘[1] 马媛[3] 毛培宏[1] 虞龙[2] 应汉杰[2] 

机构地区：[1]新疆大学物理科学与技术学院核技术及其应用技术中心,新疆乌鲁木齐830008 [2]南京工业大学生物与制药工程学院,江苏南京210009 [3]新疆大学资源与环境科学学院绿洲生态教育部重点实验室,新疆乌鲁木齐830046

出　　处：《生物技术》2010年第4期6-9,共4页Biotechnology

基　　金：国家自然科学基金项目(30760009);新疆大学自然科学基金项目(QN070109)资助

摘　　要：目的:离子注入枯草芽孢杆菌筛选高产内切葡聚糖酶突变菌株,同时进行其酶活性研究,并克隆该基因,研究离子注入对其诱变效应。方法:低能氮离子重复注入枯草芽孢杆菌,筛选获得1株高产内切葡聚糖酶突变菌株Bac11。DNS法测定酶活性。PCR扩增获得出发菌株Bac01和突变菌株Bac11内切葡聚糖酶基因,并对核酸序列及预测氨基酸序列进行多重比对。结果:突变菌株Bac11内切葡聚糖酶活性从93.33IU提高到381.89IU。多重比对Bac01和Bac11内切葡聚糖酶基因编码区1500bp序列,当中有10个碱基发生突变,预测氨基酸序列中有5个氨基酸残基发生变化,且都在其基因纤维素结合域部分。结论:低能氮离子重复注入对枯草芽孢杆菌内切葡聚糖酶活性及其基因有明显的诱变累加效应。Objective：The mutant strain with high yield of endoglucanase was isolated using nitrogen ion as mutagenic source to detect the effects of endoglucanase activity and gene mutation.Method：Bacillus subtilis was mutated repeatedly by nitrogen ion and the mutant strain Bac11with high yield of endoglucanase was screened.The endoglucanase activitiy was determined according to the DNS method,and the endoglucanase gene fragments were obtained with PCR amplification from B.subtilis Bac01 and mutant strain Bac11,and the two 1500 bp fragments and predicted amino acid sequences were analyzed with multiple alignment method.Result：It exhibited higher endoglucanase activitiy （ 381.89IU） than the original strain Bac01 （ 93.33IU） .Multiple alignment result indicated that 10 nucleotides mu-tated.Bioinformatics methods were used to analyze the two predicted amino acid sequences,and it was found that 5 amino acid residues changed,being all in the cellulose-binding domain of endoglucanase.Conclusion：The nitrogen ion irradiation on Bacillus subtilis,repeatedly,has the accumulative mutation effect on endoglucanase activity and its gene.

关 键 词：离子注入 枯草芽孢杆菌 内切葡聚糖酶 活性 基因突变 

分 类 号：Q319[生物学—遗传学] O539[理学—等离子体物理]