【摘要】 用合成生物学和代谢工程技术,筛选并克隆丁醇合成途径中酶活性较高的atoB,hbd,crt,ter,adhE2等5个关键酶基因,先通过重组PCR构建pUC18-ato B-hbd-crt-ter质粒,再将串联基因atoB-hbd-crt-ter和基因adhE2克隆至质粒pET-21b中,构建含丁醇合成途径的表达载体p ET-21b-ato B-hbd-crt-ter-adh E2,将其转入大肠杆菌E.coli BL21(DE3)中制备工程菌株.对样品进行IPTG诱导表达检测与色谱分析的结果表明,已合成出关键酶蛋白并产生一定量的丁醇.更多还原

【Abstract】 Using synthetic biology and metabolic engineering techniques,we screened and cloned 5 key enzyme gene ato B,hbd,crt,adhE2 and ter with relatively high activity in butanol synthesis pathway. Firstly,we constructed the pUC18-ato B-hbd-crt-ter plasmid by recombinant PCR,and then coloned tandem gene ato Bhbd-crt-ter and gene adhE2 into the plasmid pET-21 b and eventually constructed the expression vector of pET-21b-ato B-hbd-crt-ter-adhE2 containing butanol synthesis pathway,and transferred it into E.coli BL21( DE3)for the preparation of engineering strains. We performed IPTG induced expression detection and gas chromatograph( GC) analysis of samples. The results show that key enzyme proteins have been synthesized and produced a certain amount of butanol.更多还原