枯草杆菌纤溶酶基因的克隆及表达

为了提高豆豉纤溶酶(DFE)的表达产量,采用PCR方法从高纤溶活性的枯草芽孢杆菌DC12的基因组中扩增获得DFE基因,将含有启动子至3非翻译区的全长1400 bp基因插入大肠杆菌-枯草杆菌穿梭载体pBE3,构建了DFE基因的表达质粒,化学转化枯草杆菌WB800获得了DFE的重组表达菌.试验结果表明:DFE基因在自身启动子驱动下,在蛋白酶缺陷型的枯草杆菌中获得了高活性的分泌表达;重组表达菌株培养30 h后,培养物上清液中的纤溶酶活性最高可达690U/mL.

Cloning and Expression of Douchi Fibrinolytic Enzyme (DFE) Gene from Bacillus subtilis

In order to improve the yield of Douchi fibrinolytic enzyme(DFE),the full DFE gene encoding was amplified and cloned from the chromosome of Bacillus subtilis (B.subtilis) DC12 by means of PCR.The full DFE gene including the promoter,the encoding sequence and the 3'UTR was then inserted into the Escherichia coliB.subtilis shuttle vector pBE3 and chemically transformed into B.subtilis WB800 to construct the recombinant expression strain.The results show that DFE gene is successfully expressed under the driving of its own promoter in B.subtilis WB800 and secreted into the medium,and that,after the cultivation of recombinant strain for 30 h,the activity of fibrinolytic enzyme in the supernatant is as high as 690 U/mL.