作　　者： ; ; ; ; ;

机构地区： 广西大学

出　　处： 《生物技术通报》 2011年第11期154-159,共6页

摘　　要： 将已克隆的碱性α-淀粉酶基因信号肽编码序列去除,用PCR的方法加入酶切位点,然后与表达载体pHIS1525连接转化大肠杆菌DH5α,筛选出阳性转化子DH5α-pHIS1525-JH,并提取质粒进一步转化巨大芽孢杆菌YYBm1原生质体,获得基因工程菌YYBm1-pHIS1525-JH。SDS-PAGE分析表明该基因在巨大芽孢杆菌中得到了有效表达。酶学性质研究表明,该酶的最适温度与pH值分别为60℃与pH8.5,在pH7.0-10.5之间具有较好的稳定性,Km值为1.94 mg/mL,酶活力可达2 516.5 U。 The signal peptide coding sequence of cloned gene alkaline α-amylase was removed,with the PCR method of adding restriction sites,and then connected with the expression vector pHIS1525 transformed into E.coli DH5α,screening positive recombinant DH5α-pHIS1525-JH.The plasmid from the recombinant DH5α-pHIS1525-JH was transformed into protoplasts of Bacillus megaterium YYBm1 to obtain genetically engineered bacteria YYBm1-pHIS1525-JH.SDS-PAGE analysis showed that the gene in Bacillus megaterium haven been effective expressed.Enzyme properties studies have shown that,the optimum temperature and pH values were 60℃ and pH8.5,the enzyme is stable at pH7.0-10.5,Km value is 1.94 mg/mL,and enzyme activity was 2 516.5 U.

关 键 词： 碱性 淀粉酶 巨大芽孢杆菌 芽孢杆菌表达系统 基因表达 酶学性质

领　　域： [生物学]