具纤溶活性菌株ND2的筛选及其纳豆激酶基因的克隆、表达  

Screening of Fibrinolytic Enzyme-Producing Strain ND2 and Clone and Expression for the Natto Kinase Gene

作　　者：胡忠[1] 吴奕瑞[1] 林键[1] 庄东红[1] 

机构地区：[1]汕头大学生物学系,广东汕头515063

出　　处：《食品科学》2006年第12期473-478,共6页Food Science

摘　　要：从市售的纳豆制品中分离并纯化出一株具有高效纤溶活性的菌株ND2,经生理生化特性及16SrRNA序列分析鉴定为芽孢杆菌属的枯草芽孢杆菌。通过正交试验优化菌株ND2的发酵条件,得到ND2产纳豆激酶的最佳条件:氮源-大豆蛋白胨,碳源-淀粉,碳氮比3:1,37℃,pH6.0。经PCR扩增得到825bp的纳豆激酶成熟肽基因,采用pET32a(+)表达载体和BL21(DE3)为宿主菌,在温度为37℃、1‰IPTG浓度下可诱导获得较高表达量的重组纳豆激酶。Strain ND2 with the highest activity of the enzyme was screened from the sale natto, and was characterized as Bacillus subtilis sp. by physiologic and biochemical characteristics and analysis of 16SrRNA sequences. The optimal condition for natto kinase （NK）-fermenting was determined as follow： soybean as nitrogen source, feeula as carbon source, carbon-nitrogen ratio 3：1, 37℃, and pH6.0. The NK gene （825bp） was amplified from strain ND2 through PCR reaction. And SDS PAGE analysis revealed that using the pET32a（＋） as expression vector and BL21（DE3） as host strain and cultivating at 37℃ with 1%o IPTG, recombinant NK protein was highly expressed.

关 键 词：枯草芽孢杆菌 纳豆激酶 基因克隆与表达 

分 类 号：Q93-33[生物学—微生物学]