大豆过氧化物酶基因随机突变文库的构建

Construction of Soybean Peroxidase Gene Random Mutation Library

为提高大豆过氧化物酶(soybean peroxidase,SBP)的活性及研究该酶一级结构与酶活性间的关系,采用连续易错PCR方法对大豆过氧化物酶基因(sbp)进行了2轮随机突变,将第二轮易错PCR产物与质粒载体pPICZα-A分别进行双酶切,经T4DNA连接酶催化连接后,CaCl2法转化重组质粒pPICZα-A-sbp至大肠杆菌DH5α中,成功构建出sbp随机突变文库,库容量约为1.77×106 cfu。

In order to improve enzymatic activity of soybean peroxidase（SBP）and study the correlation between its primary structure and enzymatic activity,sequential error-prone PCR method was used on soybean peroxidase genes（sbp） for two times randon mutation.The second round error-prone PCR products and plasmid vectors pPICZα-A were digested by double enzymes.After the PCR products ligated into the vectors by T4DNA ligase,recombinant plasmids pPICZα-A-sbp were transformed into Escherichia coli DH5α through calcium chlorid method.A random mutation library was constructed,and the capacity was about 1.77×106 cfu.