【摘要】 以E.coli噬菌体λ EMBL 3为载体,用鸟枪法将地衣形芽孢杆菌的热稳定α-淀粉酶基因克隆到λ噬菌体的基因组中。携带α-淀粉酶基因的杂种噬菌体λ pAmy_αL16的DNA,经限制性内切酶HindⅢ水解后,被亚克隆到枯草杆菌的质粒pNQ 122上,并得到了表达。通过重转化作用和物理图谱分析,证明α-淀粉酶基因位于3.9 kb的Hin dⅢ DNA限制片段上。 转化子枯草杆菌(pAmy_αL41)产生的α-淀粉酶的热稳定性、最适反应温度等与亲本菌株一致。α-淀粉酶的分子量和等电点也与原菌株相同。更多还原

【Abstract】 The thermostable a-amylase gene of B.licheniformis has been cloned in E.coli using phage λEMBL3 as a vector.The HindIII restriction fragment of hybrid phage has been subcloned in B.subtilis plasmid pNQ 122.By retransformation and retriction mapping,we demonstrate that the gene coding for a-amylase is situated in 3.9 kb Hindlll restriction fragment.The expression level of the gene in B.subtilis is about 30-fold higher than the donor strain.The a-amylase produced by B.subtilis (pAmya L41) retained the thermostability,optimal reaction temprature of the enzyme produced by B.licheniformis.Its molecular weight is 60,000-62,000 doltons.The pls are 6.65 and 6.80.These results indicate that the a-amylase gene express faithfully in B.subtilis.The thermostable property of the enzyme is due to the structure of the enzyme itself.更多还原